The last universal common ancestor or last universal cellular ancestor (LUCA), also called the last universal ancestor (LUA), is the most recent population of organisms from which all organisms now living on Earth have a common descent—the most recent common ancestor of all current life on Earth. A related concept is that of progenote. LUCA is not thought to be the first life on Earth, but rather the only type of organism of its time to still have living descendants.
While there is no specific fossil evidence of LUCA, it can be studied by comparing the genomes of all modern organisms, its descendants. By these means, a 2016 study identified a set of 355 genes most likely to have been present in LUCA. The genes describe a complex life form with many co-adapted features, including transcription and translation mechanisms to convert information from DNA to RNA to proteins. The study concluded that the LUCA probably lived in the high-temperature water of deep sea vents near ocean-floor magma flows.
Studies from 2000 to 2018 have suggested an increasingly ancient
time for LUCA. In 2000, estimations suggested LUCA existed 3.5 to
3.8 billion years ago in the Paleoarcheanera, a few hundred million years before the earliest fossil evidence of life, for which there are several candidates ranging in age from 3.48 to 4.28 billion years ago. A 2018 study from the University of Bristol, applying a molecular clock model, places the LUCA shortly after 4.5 billion years ago, within the Hadean.
Charles Darwin first proposed the theory of universal common descent through an evolutionary process in his book On the Origin of Species
in 1859: "Therefore I should infer from analogy that probably all the
organic beings which have ever lived on this earth have descended from
some one primordial form, into which life was first breathed." Later biologists have separated the problem of the origin of life from that of the LUCA.
Features
By analysis of the presumed LUCA's offspring groups, the LUCA appears to have been a small, single-celled organism. It likely had a ring-shaped coil of DNA
floating freely within the cell. Morphologically, it would likely not
have stood out within a mixed population of small modern-day bacteria.
However, Carl Woeseet al., who first proposed the currently-used three domain system based on an analysis of ribosomal RNA
(rRNA) sequences of bacteria, archaea, and eukaryotes, stated that in
its genetic machinery, the LUCA would have been a "... simpler, more
rudimentary entity than the individual ancestors that spawned the three
[domains] (and their descendants)".
While the gross anatomy of LUCA can only be reconstructed with
much uncertainty, its biochemical mechanisms can be described in some
detail, based on the properties currently shared by all independently
living organisms on Earth.
Its genetic code was likely based on DNA, so that it lived after the RNA world. If DNA was present, it was composed exclusively of the four modern-day nucleotides: deoxyadenosine, deoxycytidine, deoxythymidine, and deoxyguanosine. The DNA was kept double-stranded by a template-dependent enzyme, DNA polymerase, which was recently proposed to belong to the family D. The integrity of the DNA benefited from a group of maintenance and repair enzymes including DNA topoisomerase.
If the genetic code was DNA-based, it was expressed via single-stranded
RNA intermediates. The RNA was produced by a DNA-dependent RNA polymerase using nucleotides similar to those of DNA, with the exception that the DNA nucleotide thymidine was replaced by uridine in RNA. It had multiple DNA-binding proteins, such as histone-fold proteins.
The genetic code was expressed into proteins. These were assembled from free amino acids by translation of a messenger RNA via a mechanism of ribosomes, transfer RNAs, and a group of related proteins. The ribosomes were composed of two subunits, a big 50S and a small 30S. Each ribosomal subunit was composed of a core of ribosomal RNA surrounded by ribosomal proteins.
Both types of RNA molecules (ribosomal and transfer RNAs) played an
important role in the catalytic activity of the ribosomes. Only 20 amino acids were used, only in L-isomers, to the exclusion of countless other amino acids. ATP served as an energy intermediate. Several hundred protein enzymes
catalyzed chemical reactions to extract energy from fats, sugars, and
amino acids, and to synthesize fats, sugars, amino acids, and nucleic
acid bases through various chemical pathways.
The cell contained a water-based cytoplasm effectively enclosed by a lipid bilayer membrane. The cell tended to exclude sodium and concentrate potassium by means of specific ion transporters (or ion pumps). The cell multiplied by duplicating all its contents followed by cellular division. The cell used chemiosmosis to produce energy. It also reduced CO2 and oxidized H2 (methanogenesis or acetogenesis) via acetyl-thioesters.
A 1990 phylogenetic tree linking all major groups of living organisms to the LUCA (the black trunk at the bottom), based on ribosomal RNA sequence data.
In 1859, Charles Darwin published On the Origin of Species, in which he twice stated the hypothesis that there was only one progenitor for all life forms. In the summation he states:
"Therefore I should infer from analogy that probably all the
organic beings which have ever lived on this earth have descended from
some one primordial form, into which life was first breathed."
The last sentence begins with a restatement of the hypothesis:
"There is grandeur in this view of life, with its several
powers, having been originally breathed into a few forms or into
one ..."
When the LUCA was hypothesized, cladograms based on genetic distance between living cells indicated that Archaea
split early from the rest of living things. This was inferred from the
fact that the archaeans known at that time were highly resistant to
environmental extremes such as high salinity, temperature or acidity, leading some scientists to suggest that the LUCA evolved in areas like the deep ocean vents,
where such extremes prevail today. Archaea, however, were later
discovered in less hostile environments, and are now believed to be more
closely related to the Eukaryota than to the Bacteria, although many details are still unknown.
In 2010, based on "the vast array of molecular sequences now available from all domains of life," a formal test of universal common ancestry was published. The formal test favored the existence of a universal common ancestor over a wide class of alternative hypotheses that included horizontal gene transfer.
Basic biochemical principles make it overwhelmingly likely that all
organisms do have a single common ancestor. It is extremely unlikely
that organisms descended from separate incidents of cell-formation would
be able to complete a horizontal gene transfer without garbling each
other's genes, converting them into noncoding segments.
Further, many more amino acids are chemically possible than the 22
found in protein molecules. These lines of chemical evidence,
incorporated into the formal statistical test point to a single cell
having been the LUCA. While the test overwhelmingly favored the
existence of a single LUCA, this does not imply that the LUCA was ever
alone: Instead, it was one of many early microbes but the only one whose descendants survived beyond the Paleoarchean Era.
With the later gene pool of the LUCA's descendants, with their common framework of the AT/GC rule and the standard twenty amino acids, horizontal gene transfer would have been feasible and could have been very common.
In an earlier hypothesis, Carl Woese (1988) had proposed that:
no individual organism can be considered a LUCA, and
the genetic heritage of all modern organisms derived through horizontal gene transfer among an ancient community of organisms.
While the results of Theobald (2010) and Saey (2010) demonstrated the
existence of a single LUCA, Woese's argument can still be applied to
Ur-organisms (initial products of abiogenesis) before the LUCA. At the
beginnings of life, ancestry was not as linear as it is today because
the genetic code had not evolved.
Before high fidelity replication, organisms could not be easily mapped
on a phylogenetic tree. However, the LUCA lived after the origin of the
genetic code and at least some rudimentary early form of molecular
proofreading.
The most commonly accepted tree of life, based on several molecular studies, has its root between a monophyleticdomainBacteria and a clade formed by Archaea and Eukaryota. However, a very small minority of studies place the root in the domain Bacteria, in the phylum Firmicutes, or state that the phylum Chloroflexi is basal to a clade with Archaea and Eukaryotes and the rest of Bacteria (as proposed by Thomas Cavalier-Smith).
Research by William F. Martin
(2016) genetically analyzed 6.1 million protein-coding genes and
286,514 protein clusters from sequenced prokaryotic genomes of various
phylogenetic trees, and identified 355 protein clusters that were
probably common to the LUCA. The results "depict LUCA as anaerobic, CO2-fixing, H2-dependent with a Wood–Ljungdahl pathway (the reductive acetyl-coenzyme A pathway), N2-fixing and thermophilic. LUCA's biochemistry was replete with FeS clusters and radical reaction mechanisms." The cofactors also reveal "dependence upon transition metals, flavins, S-adenosyl methionine, coenzyme A, ferredoxin, molybdopterin, corrins and selenium. Its genetic code required nucleoside modifications and S-adenosylmethionine-dependent methylations." The results are "quite specific": they show that methanogenicclostridia was a basal clade in the 355 lineages examined, and that the LUCA may therefore have inhabited an anaerobic hydrothermal vent setting in a geochemically active environment rich in H2, CO2, and iron.
These findings could mean that life on Earth originated in such
hydrothermal vents, but it is also possible that life was restricted to
such locations at some later time, perhaps by the Late Heavy Bombardment.
The identification of these genes as being present in LUCA has also
been criticized, as they may simply represent later genes which migrated
via horizontal gene transfers between archaea and bacteria.
Viruses
Based on the extant distribution of viruses across the two primary domains of life, bacteria and archaea,
it has been suggested that LUCA was associated with a remarkably
complex virome that already included the main groups of extant viruses
of bacteria and archaea and that extensive virus evolution has
antedated, or preceded in time, the LUCA. This ancestral virome was likely dominated by dsDNA viruses from the realms Duplodnaviria and Varidnaviria. In addition, two groups of single-stranded DNA viruses (realm Monodnaviria), namely Microviridae and Tubulavirales,
can be traced to the last bacterial common ancestor (LBCA), whereas
spindle-shaped viruses most likely infected the last archaeal common
ancestor (LACA). The possibility that these virus groups were present in
the LUCA virome but were subsequently lost in one of the two primary
domains cannot be dismissed. By contrast, RNA viruses do not appear to
have been a prominent part of the LUCA virome, even though
straightforward thinking might have envisaged the LUCA virome as a
domain of RNA viruses descending from the primordial RNA world. Instead,
by the time the LUCA lived, RNA viruses had probably already been
largely supplanted by the more efficient DNA virosphere.
RNA-based evolution is a theory that posits that RNA is not merely an intermediate between Watson and Crick model of the DNA molecule and proteins, but rather a far more dynamic and independent role-player in determining phenotype. By regulating the transcription in DNA sequences, the stability of RNA, and the capability of messenger RNA to be translated, RNA processing events allow for a diverse array of proteins to be synthesized from a single gene. Since RNA processing is heritable, it is subject to natural selection suggested by Darwin and contributes to the evolution and diversity of most eukaryotic organisms.
Role of RNA in conventional evolution
In accordance with the central dogma of molecular biology, RNA passes information between the DNA of a genome and the proteins expressed within an organism. Therefore, from an evolutionary standpoint, a mutation
within the DNA bases results in an alteration of the RNA transcripts,
which in turn leads to a direct difference in phenotype.
RNA is also believed to have been the genetic material of the first life
on Earth. The role of RNA in the origin of life is best supported by
the ease of forming RNA from basic chemical building blocks (such as amino acids, sugars, and hydroxyl acids) that were likely present 4 billion years ago. Molecules of RNA have also been shown to effectively self-replicate, catalyze basic reactions, and store heritable information.
As life progressed and evolved over time only DNA, which is much more
chemically stable than RNA, could support large genomes and eventually
took over the role as the major carrier of genetic information.
Variability of RNA processing
Research
within the past decade has shown that strands of RNA are not merely
transcribed from regions of DNA and translated into proteins. Rather
RNA has retained some of its former independence from DNA, and is
subject to a network of processing events that alter the protein
expression from that bounded by just the genomic DNA.
Processing of RNA influences protein expression by managing the
transcription of DNA sequences, the stability of RNA, and the
translation of messenger RNA.
Alternative splicing
Splicing
is the process by which non-coding regions of RNA are removed. The
number and combination of splicing events varies greatly based on
differences in transcript sequence and environmental factors. Variation
in phenotype caused by alternative splicing is best seen in the sex
determination of D. melanogaster. The Tra gene, determinant of sex, in male flies becomes truncated as splicing events fail to remove a stop codon
that controls the length of the RNA molecule. In others the stop
signal is retained within the final RNA molecule and a functional Tra
protein is produced resulting in the female phenotype. Thus, alternative RNA splicing events allow differential phenotypes, regardless of the identity of the coding DNA sequence.
RNA stability
Phenotype
may also be determined by the number of RNA molecules, as more RNA
transcripts lead to a greater expression of protein. Short tails of
repetitive nucleic acids are often added to the ends of RNA molecules in
order to prevent degradation, effectively increasing the number of RNA
strands able to be translated into protein. During mammalian liver regeneration RNA molecules of growth factors increase in number due to the addition of signaling tails. With more transcripts present the growth factors are produced at a higher rate, aiding the rebuilding process of the organ.
RNA silencing
Silencing
of RNA occurs when double stranded RNA molecules are processed by a
series of enzymatic reactions, resulting in RNA fragments that degrade
complementary RNA sequences.
By degrading transcripts, a lower amount of protein products are
translated and the phenotype is altered by yet another RNA processing
event.
Evolutionary mechanism
Most
RNA processing events work in concert with one another and produce
networks of regulating processes that allow a greater variety of
proteins to be expressed than those strictly directed by the genome. These RNA processing events can also be passed on from generation to generation via reverse transcription into the genome.
Over time, RNA networks that produce the fittest phenotypes will be
more likely to be maintained in a population, contributing to evolution.
Studies have shown that RNA processing events have especially been
critical with the fast phenotypic evolution of vertebrates—large jumps in phenotype explained by changes in RNA processing events.
Human genome searches have also revealed RNA processing events that
have provided significant “sequence space for more variability”.
On the whole, RNA processing expands the possible phenotypes of a
given genotype and contributes to the evolution and diversity of life.
RNA virus evolution
RNA
virus evolution appears to be facilitated by a high mutation rate
caused by the lack of a proofreading mechanism during viral genome
replication. In addition to mutation, RNA virus evolution is also facilitated by genetic recombination. Genetic recombination can occur when at least two RNA viral genomes are present in the same host cell, and has been studies in numerous RNA viruses. RNA recombination appears to be a major driving force in viral evolution among Picornaviridae ((+)ssRNA) (e.g. poliovirus). In the Retroviridae ((+)ssRNA)(e.g. HIV), damage in the RNA genome appears to be avoided during reverse transcription by strand switching, a form of genetic recombination. Recombination also occurs in the Coronaviridae ((+)ssRNA) (e.g. SARS). Recombination in RNA viruses appears to be an adaptation for coping with genome damage.
Recombination can occur infrequently between animal viruses of the
same species but of divergent lineages. The resulting recombinant
viruses may sometimes cause an outbreak of infection in humans.
Tumor protein P53, also known as p53, cellular tumor antigen p53 (UniProt name), the Guardian of the Genome, phosphoprotein p53, tumor suppressor p53, antigen NY-CO-13, or transformation-related protein 53 (TRP53), is any isoform of a protein encoded by homologous genes in various organisms, such as TP53 (humans) and Trp53 (mice). This homolog (originally thought to be, and often spoken of as, a single protein) is crucial in multicellularvertebrates, where it prevents cancer formation, and thus functions as a tumor suppressor. As such, p53 has been described as "the guardian of the genome" because of its role in conserving stability by preventing genome mutation. Hence TP53 is classified as a tumor suppressor gene.
The name p53 was given in 1979 describing the apparent molecular mass; SDS-PAGE analysis indicates that it is a 53-kilodalton (kDa) protein. However, the actual mass of the full-length p53 protein (p53α) based on the sum of masses of the amino acid residues is only 43.7 kDa. This difference is due to the high number of proline residues in the protein, which slow its migration on SDS-PAGE, thus making it appear heavier than it actually is. In addition to the full-length protein, the human TP53 gene encodes at least 15 protein isoforms, ranging in size from 3.5 to 43.7 kDa. All these p53 proteins are called the p53 isoforms. The TP53 gene is the most frequently mutated gene (>50%) in human cancer, indicating that the TP53 gene plays a crucial role in preventing cancer formation. TP53 gene encodes proteins that bind to DNA and regulate gene expression to prevent mutations of the genome.
Gene
In humans, the TP53 gene is located on the short arm of chromosome 17 (17p13.1).
The gene spans 20 kb, with a non-coding exon 1 and a very long first
intron of 10 kb. The coding sequence contains five regions showing a
high degree of conservation in vertebrates, predominantly in exons 2, 5,
6, 7 and 8, but the sequences found in invertebrates show only distant
resemblance to mammalian TP53. TP53orthologs have been identified in most mammals for which complete genome data are available.
Human TP53 gene
In humans, a common polymorphism involves the substitution of an arginine for a proline at codon
position 72. Many studies have investigated a genetic link between this
variation and cancer susceptibility; however, the results have been
controversial. For instance, a meta-analysis from 2009 failed to show a
link for cervical cancer. A 2011 study found that the TP53 proline mutation did have a profound effect on pancreatic cancer risk among males. A study of Arab women found that proline homozygosity at TP53 codon 72 is associated with a decreased risk for breast cancer. One study suggested that TP53 codon 72 polymorphisms, MDM2 SNP309, and A2164G may collectively be associated with non-oropharyngeal cancer susceptibility and that MDM2 SNP309 in combination with TP53 codon 72 may accelerate the development of non-oropharyngeal cancer in women. A 2011 study found that TP53 codon 72 polymorphism was associated with an increased risk of lung cancer.
Meta-analyses from 2011 found no significant associations between TP53 codon 72 polymorphisms and both colorectal cancer risk and endometrial cancer risk. A 2011 study of a Brazilian birth cohort found an association between the non-mutant arginine TP53 and individuals without a family history of cancer.
Another 2011 study found that the p53 homozygous (Pro/Pro) genotype was
associated with a significantly increased risk for renal cell
carcinoma.
Structure
A schematic of the known protein domains in p53. (NLS = Nuclear Localization Signal).
Crystal structure of four p53 DNA binding domains (as found in the bioactive homo-tetramer) and has seven domains:
an acidic N-terminus transcription-activation domain (TAD), also known as activation domain 1 (AD1), which activates transcription factors.
The N-terminus contains two complementary transcriptional activation
domains, with a major one at residues 1–42 and a minor one at residues
55–75, specifically involved in the regulation of several pro-apoptotic
genes.
activation domain 2 (AD2) important for apoptotic activity: residues 43–63.
proline rich domain important for the apoptotic activity of p53 by nuclear exportation via MAPK: residues 64–92.
central DNA-binding core domain (DBD). Contains one zinc atom and several arginine amino acids: residues 102–292. This region is responsible for binding the p53 co-repressor LMO3.
homo-oligomerisation domain (OD): residues 307–355. Tetramerization is essential for the activity of p53 in vivo.
C-terminal involved in downregulation of DNA binding of the central domain: residues 356–393.
Mutations that deactivate p53 in cancer usually occur in the DBD.
Most of these mutations destroy the ability of the protein to bind to
its target DNA sequences, and thus prevents transcriptional activation
of these genes. As such, mutations in the DBD are recessiveloss-of-function mutations. Molecules of p53 with mutations in the OD dimerise with wild-type
p53, and prevent them from activating transcription. Therefore, OD
mutations have a dominant negative effect on the function of p53.
p53 plays a role in regulation or progression through the cell cycle, apoptosis, and genomic stability by means of several mechanisms:
It can activate DNA repair proteins when DNA has sustained damage. Thus, it may be an important factor in aging.
It can arrest growth by holding the cell cycle at the G1/S regulation point
on DNA damage recognition—if it holds the cell here for long enough,
the DNA repair proteins will have time to fix the damage and the cell
will be allowed to continue the cell cycle.
It can initiate apoptosis (i.e., programmed cell death) if DNA damage proves to be irreparable.
p53 pathway:
In a normal cell, p53 is inactivated by its negative regulator, mdm2.
Upon DNA damage or other stresses, various pathways will lead to the
dissociation of the p53 and mdm2 complex. Once activated, p53 will
induce a cell cycle arrest to allow either repair and survival of the
cell or apoptosis to discard the damaged cell. How p53 makes this choice
is currently unknown.
WAF1/CIP1 encoding for p21 and hundreds of other down-stream genes. p21 (WAF1) binds to the G1-S/CDK (CDK4/CDK6, CDK2, and CDK1) complexes (molecules important for the G1/S transition in the cell cycle) inhibiting their activity.
When p21(WAF1) is complexed with CDK2, the cell cannot continue
to the next stage of cell division. A mutant p53 will no longer bind DNA
in an effective way, and, as a consequence, the p21 protein will not be
available to act as the "stop signal" for cell division.
Studies of human embryonic stem cells (hESCs) commonly describe the
nonfunctional p53-p21 axis of the G1/S checkpoint pathway with
subsequent relevance for cell cycle regulation and the DNA damage
response (DDR). Importantly, p21 mRNA is clearly present and upregulated
after the DDR in hESCs, but p21 protein is not detectable. In this cell
type, p53 activates numerous microRNAs (like miR-302a, miR-302b, miR-302c, and miR-302d) that directly inhibit the p21 expression in hESCs.
The p21 protein binds directly to cyclin-CDK complexes that drive
forward the cell cycle and inhibits their kinase activity, thereby
causing cell cycle arrest to allow repair to take place. p21 can also
mediate growth arrest associated with differentiation and a more
permanent growth arrest associated with cellular senescence. The p21
gene contains several p53 response elements that mediate direct binding
of the p53 protein, resulting in transcriptional activation of the gene
encoding the p21 protein.
The p53 and RB1 pathways are linked via p14ARF, raising the possibility that the pathways may regulate each other.
p53 expression can be stimulated by UV light, which also causes DNA damage. In this case, p53 can initiate events leading to tanning.
Stem cells
Levels of p53 play an important role in the maintenance of stem cells throughout development and the rest of human life.
In human embryonic stem cells (hESCs)s, p53 is maintained at low inactive levels. This is because activation of p53 leads to rapid differentiation of hESCs.
Studies have shown that knocking out p53 delays differentiation and
that adding p53 causes spontaneous differentiation, showing how p53
promotes differentiation of hESCs and plays a key role in cell cycle as a
differentiation regulator. When p53 becomes stabilized and activated in
hESCs, it increases p21 to establish a longer G1. This typically leads
to abolition of S-phase entry, which stops the cell cycle in G1, leading
to differentiation. Work in mouse embryonic stem cells has recently
shown however that the expression of P53 does not necessarily lead to
differentiation. p53 also activates miR-34a and miR-145, which then repress the hESCs pluripotency factors, further instigating differentiation.
In adult stem cells, p53 regulation is important for maintenance of stemness in adult stem cell niches. Mechanical signals such as hypoxia affect levels of p53 in these niche cells through the hypoxia inducible factors, HIF-1α and HIF-2α. While HIF-1α stabilizes p53, HIF-2α suppresses it.
Suppression of p53 plays important roles in cancer stem cell phenotype,
induced pluripotent stem cells and other stem cell roles and behaviors,
such as blastema formation. Cells with decreased levels of p53 have
been shown to reprogram into stem cells with a much greater efficiency
than normal cells.
Papers suggest that the lack of cell cycle arrest and apoptosis gives
more cells the chance to be reprogrammed. Decreased levels of p53 were
also shown to be a crucial aspect of blastema formation in the legs of salamanders.
p53 regulation is very important in acting as a barrier between stem
cells and a differentiated stem cell state, as well as a barrier between
stem cells being functional and being cancerous.
Other
Apart from the cellular and molecular effects above, p53 has a tissue-level anticancer effect that works by inhibiting angiogenesis.
As tumors grow they need to recruit new blood vessels to supply them,
and p53 inhibits that by (i) interfering with regulators of tumor hypoxia
that also affect angiogenesis, such as HIF1 and HIF2, (ii) inhibiting
the production of angiogenic promoting factors, and (iii) directly
increasing the production of angiogenesis inhibitors, such as arresten.
p53 becomes activated in response to myriad stressors, including but not limited to DNA damage (induced by either UV, IR, or chemical agents such as hydrogen peroxide), oxidative stress, osmotic shock,
ribonucleotide depletion, and deregulated oncogene expression. This
activation is marked by two major events. First, the half-life of the
p53 protein is increased drastically, leading to a quick accumulation of
p53 in stressed cells. Second, a conformational change forces p53 to be activated as a transcription regulator
in these cells. The critical event leading to the activation of p53 is
the phosphorylation of its N-terminal domain. The N-terminal
transcriptional activation domain contains a large number of
phosphorylation sites and can be considered as the primary target for
protein kinases transducing stress signals.
The protein kinases
that are known to target this transcriptional activation domain of p53
can be roughly divided into two groups. A first group of protein kinases
belongs to the MAPK
family (JNK1-3, ERK1-2, p38 MAPK), which is known to respond to several
types of stress, such as membrane damage, oxidative stress, osmotic
shock, heat shock, etc. A second group of protein kinases (ATR, ATM, CHK1 and CHK2, DNA-PK, CAK, TP53RK)
is implicated in the genome integrity checkpoint, a molecular cascade
that detects and responds to several forms of DNA damage caused by
genotoxic stress. Oncogenes also stimulate p53 activation, mediated by
the protein p14ARF.
In unstressed cells, p53 levels are kept low through a continuous degradation of p53. A protein called Mdm2 (also called HDM2 in humans), binds to p53, preventing its action and transports it from the nucleus to the cytosol. Mdm2 also acts as an ubiquitin ligase and covalently attaches ubiquitin to p53 and thus marks p53 for degradation by the proteasome. However, ubiquitylation of p53 is reversible. On activation of p53, Mdm2 is also activated, setting up a feedback loop. p53 levels can show oscillations
(or repeated pulses) in response to certain stresses, and these pulses
can be important in determining whether the cells survive the stress, or
die.
MI-63 binds to MDM2, reactivating p53 in situations where p53's function has become inhibited.
A ubiquitin specific protease, USP7 (or HAUSP), can cleave ubiquitin off p53, thereby protecting it from proteasome-dependent degradation via the ubiquitin ligase pathway . This is one means by which p53 is stabilized in response to oncogenic insults. USP42 has also been shown to deubiquitinate p53 and may be required for the ability of p53 to respond to stress.
Recent research has shown that HAUSP is mainly localized in the
nucleus, though a fraction of it can be found in the cytoplasm and
mitochondria. Overexpression of HAUSP results in p53 stabilization.
However, depletion of HAUSP does not result to a decrease in p53 levels
but rather increases p53 levels due to the fact that HAUSP binds and
deubiquitinates Mdm2. It has been shown that HAUSP is a better binding
partner to Mdm2 than p53 in unstressed cells.
USP10 however has been shown to be located in the cytoplasm in
unstressed cells and deubiquitinates cytoplasmic p53, reversing Mdm2
ubiquitination. Following DNA damage, USP10 translocates to the nucleus
and contributes to p53 stability. Also USP10 does not interact with
Mdm2.
Phosphorylation of the N-terminal end of p53 by the
above-mentioned protein kinases disrupts Mdm2-binding. Other proteins,
such as Pin1, are then recruited to p53 and induce a conformational
change in p53, which prevents Mdm2-binding even more. Phosphorylation
also allows for binding of transcriptional coactivators, like p300 and PCAF,
which then acetylate the carboxy-terminal end of p53, exposing the DNA
binding domain of p53, allowing it to activate or repress specific
genes. Deacetylase enzymes, such as Sirt1 and Sirt7, can deacetylate p53, leading to an inhibition of apoptosis. Some oncogenes can also stimulate the transcription of proteins that bind to MDM2 and inhibit its activity.
Role in disease
Overview of signal transduction pathways involved in apoptosis.
If the TP53 gene is damaged, tumor suppression is severely compromised. People who inherit only one functional copy of the TP53 gene will most likely develop tumors in early adulthood, a disorder known as Li-Fraumeni syndrome.
The TP53 gene can also be modified by mutagens (chemicals, radiation, or viruses), increasing the likelihood for uncontrolled cell division. More than 50 percent of human tumors contain a mutation or deletion of the TP53 gene. Loss of p53 creates genomic instability that most often results in an aneuploidy phenotype.
Increasing the amount of p53 may seem a solution for treatment of
tumors or prevention of their spreading. This, however, is not a usable
method of treatment, since it can cause premature aging. Restoring endogenous
normal p53 function holds some promise. Research has shown that this
restoration can lead to regression of certain cancer cells without
damaging other cells in the process. The ways by which tumor regression
occurs depends mainly on the tumor type. For example, restoration of
endogenous p53 function in lymphomas may induce apoptosis,
while cell growth may be reduced to normal levels. Thus,
pharmacological reactivation of p53 presents itself as a viable cancer
treatment option. The first commercial gene therapy, Gendicine, was approved in China in 2003 for the treatment of head and neck squamous cell carcinoma. It delivers a functional copy of the p53 gene using an engineered adenovirus.
Certain pathogens can also affect the p53 protein that the TP53 gene expresses. One such example, human papillomavirus
(HPV), encodes a protein, E6, which binds to the p53 protein and
inactivates it. This mechanism, in synergy with the inactivation of the
cell cycle regulator pRb by the HPV protein E7, allows for repeated cell division manifested clinically as warts. Certain HPV types, in particular types 16 and 18, can also lead to progression from a benign wart to low or high-grade cervical dysplasia, which are reversible forms of precancerous lesions. Persistent infection of the cervix over the years can cause irreversible changes leading to carcinoma in situ
and eventually invasive cervical cancer. This results from the effects
of HPV genes, particularly those encoding E6 and E7, which are the two
viral oncoproteins that are preferentially retained and expressed in
cervical cancers by integration of the viral DNA into the host genome.
The p53 protein is continually produced and degraded in cells of
healthy people, resulting in damped oscillation. The degradation of the
p53 protein is associated with binding of MDM2. In a negative feedback
loop, MDM2 itself is induced by the p53 protein. Mutant p53 proteins
often fail to induce MDM2, causing p53 to accumulate at very high
levels. Moreover, the mutant p53 protein itself can inhibit normal p53
protein levels. In some cases, single missense mutations in p53 have
been shown to disrupt p53 stability and function.
Suppression of p53 in human breast cancer cells is shown to lead to increased CXCR5 chemokine receptor gene expression and activated cell migration in response to chemokineCXCL13.
One study found that p53 and Myc proteins were key to the survival of Chronic Myeloid Leukaemia (CML) cells. Targeting p53 and Myc proteins with drugs gave positive results on mice with CML.
Experimental analysis of p53 mutations
Most
p53 mutations are detected by DNA sequencing. However, it is known that
single missense mutations can have a large spectrum from rather mild to
very severe functional affects.
The large spectrum of cancer phenotypes due to mutations in the TP53 gene is also supported by the fact that different isoforms of p53 proteins have different cellular mechanisms for prevention against cancer. Mutations in TP53
can give rise to different isoforms, preventing their overall
functionality in different cellular mechanisms and thereby extending the
cancer phenotype from mild to severe. Recents studies show that p53
isoforms are differentially expressed in different human tissues, and
the loss-of-function or gain-of-function mutations within the isoforms can cause tissue-specific cancer or provides cancer stem cellpotential in different tissues. TP53 mutation also hits energy metabolism and increases glycolysis in breast cancer cells.
The dynamics of p53 proteins, along with its antagonist Mdm2, indicate that the levels of p53, in units of concentration, oscillate as a function of time. This "damped" oscillation is both clinically documented and mathematically modelled. Mathematical models also indicate that the p53 concentration oscillates much faster once teratogens, such as double-stranded breaks (DSB) or UV radiation, are introduced to the system. This supports and models the current understanding of p53 dynamics, where DNA damage induces p53 activation (see p53 regulation
for more information). Current models can also be useful for modelling
the mutations in p53 isoforms and their effects on p53 oscillation,
thereby promoting de novo tissue-specific pharmacological drug discovery.
Warren Maltzman, of the Waksman Institute of Rutgers University
first demonstrated that TP53 was responsive to DNA damage in the form of
ultraviolet radiation. In a series of publications in 1991–92, Michael Kastan of Johns Hopkins University, reported that TP53 was a critical part of a signal transduction pathway that helped cells respond to DNA damage.
In 1993, p53 was voted molecule of the year by Science magazine.
Isoforms
As with 95% of human genes, TP53 encodes more than one protein. Several isoforms
were discovered in 2005, and so far 12 human p53 isoforms have been
identified (p53α, p53β, p53γ, ∆40p53α, ∆40p53β, ∆40p53γ, ∆133p53α,
∆133p53β, ∆133p53γ, ∆160p53α, ∆160p53β, ∆160p53γ). Furthermore, p53
isoforms are expressed in a tissue dependent manner and p53α is never
expressed alone.
The full length p53 isoform proteins can be subdivided into different protein domains.
Starting from the N-terminus, there are first the amino-terminal
transactivation domains (TAD 1, TAD 2), which are needed to induce a
subset of p53 target genes. This domain is followed by the proline rich
domain (PXXP), whereby the motif PXXP is repeated (P is a proline and X
can be any amino acid). It is required among others for p53 mediated apoptosis.
Some isoforms lack the proline rich domain, such as Δ133p53β,γ and
Δ160p53α,β,γ; hence some isoforms of p53 are not mediating apoptosis,
emphasizing the diversifying roles of the TP53 gene. Afterwards there is the DNA binding domain (DBD), which enables the proteins to sequence specific binding. The carboxyl terminal
domain completes the protein. It includes the nuclear localization
signal (NLS), the nuclear export signal (NES) and the oligomerisation
domain (OD). The NLS and NES are responsible for the subcellular
regulation of p53. Through the OD, p53 can form a tetramer and then bind
to DNA. Among the isoforms, some domains can be missing, but all of
them share most of the highly conserved DNA-binding domain.
The isoforms are formed by different mechanisms. The beta and the
gamma isoforms are generated by multiple splicing of intron 9, which
leads to a different C-terminus. Furthermore, the usage of an internal
promoter in intron 4 causes the ∆133 and ∆160 isoforms, which lack the
TAD domain and a part of the DBD. Moreover, alternative initiation of
translation at codon 40 or 160 bear the ∆40p53 and ∆160p53 isoforms.
Due to the isoformic nature of p53 proteins, there have been several sources of evidence showing that mutations within the TP53
gene giving rise to mutated isoforms are causative agents of various
cancer phenotypes, from mild to severe, due to single mutation in the TP53 gene (refer to section Experimental analysis of p53 mutations for more details).
Cells were first seen in 17th century Europe with the invention of the compound microscope. In 1665, Robert Hooke termed the building block of all living organisms as "cells" after looking at a piece of cork and observing a cell-like structure, however, the cells were dead and gave no indication to the actual overall components of a cell. A few years later, in 1674, Anton Van Leeuwenhoek was the first to analyze live cells in his examination of algae. All of this preceded the cell theory
which states that all living things are made up of cells and that cells
are the functional and structural unit of organisms. This was
ultimately concluded by plant scientist, Matthias Schleiden and animal scientist, Theodor Schwann in 1838, who viewed live cells in plant and animal tissue, respectively. 19 years later, Rudolf Virchow further contributed to the cell theory, adding that all cells come from the division of pre-existing cells.
Although widely accepted, there have been many studies that question
the validity of the cell theory. Viruses, for example, lack common
characteristics of a living cell, such as membranes, cell organelles, and the ability to reproduce by themselves. Scientists have struggled to decide whether viruses are alive or not and whether they are in agreement with the cell theory.
Techniques
Modern-day
cell biology research looks at different ways to culture and manipulate
cells outside of a living body to further research in human anatomy and
physiology, and to derive medications. The techniques by which cells
are studied have evolved. Due to advancements in microscopy, techniques
and technology have allowed for scientists to hold a better
understanding of the structure and function of cells. Many techniques
commonly used to study cell biology are listed below:
Cell culture:
Utilizes rapidly growing cells on media which allows for a large amount
of a specific cell type and an efficient way to study cells.
Fluorescence microscopy: Fluorescent markers such as GFP,
are used to label a specific component of the cell. Afterwards, a
certain light wavelength is used to excite the fluorescent marker which
can then be visualized.
Phase-contrast microscopy: Uses the optical aspect of light to represent the solid, liquid, and gas phase changes as brightness differences.
Confocal microscopy: Combines fluorescence microscopy with imaging by focusing light and snap shooting instances to form a 3-D image.
Transmission electron microscopy:
Involves metal staining and the passing of electrons through the cells,
which will be deflected upon interaction with metal. This ultimately
forms an image of the components being studied.
Cytometry:
The cells are placed in the machine which uses a beam to scatter the
cells based on different aspects and can therefore separate them based
on size and content. Cells may also be tagged with GFP-florescence and
can be separated that way as well.
Cell fractionation: This process requires breaking up the cell using high temperature or sonification followed by centrifugation to separate the parts of the cell allowing for them to be studied separately.
Cell classification and composition
There are two fundamental classifications of cells: prokaryotic and eukaryotic. Prokaryotic cells are distinguished from eukaryotic cells by the absence of a cell nucleus or other membrane bound organelle. Prokaryotic cells are much smaller than eukaryotic cells, making them the smallest form of life. The study of eukaryotic cells is typically the main focus of cytologists, whereas prokaryotic cells are the focus of microbiologists.
Flagella: A tail-like structure that helps the cell to move.
Ribosomes: Used for translation of RNA to protein.
Nucleoid: Area designated to hold all the genetic material in a circular structure.
There are many process that occur in prokaryotic cells that allow them to survive. For instance, in a process termed conjugation,
fertility factor allows the bacteria to possess a pilus which allows it
to transmit DNA to another bacteria which lacks the F factor,
permitting the transmittance of resistance allowing it to survive in
certain environments.
Eukaryotic cells
A typical animal cell.
Eukaryotic cells can either be unicellular or multicellular and include animal, plant, fungi, and protozoa cells which all contain organelles with various shapes and sizes. These cells are composed of the following organelles:
Nucleus:
This functions as the genome and genetic information storage for the
cell, containing all the DNA organized in the form of chromosomes. It is
surrounded by a nuclear envelope, which includes nuclear pores allowing for transportation of proteins between the inside and outside of the nucleus.
This is also the site for replication of DNA as well as transcription
of DNA to RNA. Afterwards, the RNA is modified and transported out to
the cytosol to be translated to protein.
Nucleolus:
This structure is within the nucleus, usually dense and spherical in
shape. It is the site of ribosomal RNA (rRNA) synthesis, which is needed
for ribosomal assembly.
Endoplasmic reticulum (ER): This functions to synthesize, store, and secrete proteins to the golgi apparatus.
Mitochondria:
This functions for the production of energy or ATP within the cell.
Specifically, this is the place where the Krebs cycle or TCA cycle for
the production of NADH and FADH occurs. Afterwards, these products are
used within the electron transport chain (ETC) and oxidative
phosphorylation for the final production of ATP.
Golgi apparatus:
This functions to further process, package, and secrete the proteins to
their destination. The proteins contain a signal sequence which allows
the golgi apparatus to recognize and direct it to the correct place.
Lysosome:
The lysosome functions to degrade material brought in from the outside
of the cell or old organelles. This contains many acid hydrolases,
proteases, nucleases, and lipases, which breakdown the various
molecules. Autophagy
is the process of degradation through lysosomes which occurs when a
vesicle buds off from the ER and engulfs the material, then, attaches
and fuses with the lysosome to allow the material to be degraded.
Cytoskeleton: This functions to anchor organelles within the cells and make up the structure and stability of the cell.
Cell membrane: The cell membrane can be described as a phospholipid bilayer and is also consisted of lipids and proteins.
Because the inside of the bilayer is hydrophobic and in order for
molecules to participate in reactions within the cell, they need to be
able to cross this membrane layer to get into cell via osmotic pressure, diffusion, concentration gradients, and membrane channels.
Centrioles: Function to produce spindle fibers which are used to separate chromosomes during cell division.
Eukaryotic cells may also be composed of the following molecular components:
Chromatin: This makes up chromosomes and is a mixture of DNA with various proteins.
Cilia : They help to propel substances and can also be used for sensory purposes.
Processes
Cell metabolism
Cell
metabolism is necessary for the production of energy for the cell and
therefore its survival and includes many pathways. For cellular respiration,
once glucose is available, glycolysis occurs within the cytosol of the
cell to produce pyruvate. Pyruvate undergoes decarboxylation using the
multi-enzyme complex to form acetyl coA which can readily be used in the
TCA cycle to produce NADH and FADH2. These products are involved in the electron transport chain
to ultimately form a proton gradient across the inner mitochondrial
membrane. This gradient can then drive the production of ATP and H2O
during oxidative phosphorylation. Metabolism in plant cells includes photosynthesis which is simply the exact opposite of respiration as it ultimately produces molecules of glucose.
Cell communication and signaling
Cell
communication is important for cell regulation and for cells to process
information from the environment and respond accordingly. Communication
can occur through direct cell contact or endocrine, paracrine, and autocrine signaling.
Direct cell-cell contact is when a receptor on a cell binds a molecule
that is attached to the membrane of another cell. Endocrine signaling
occurs through molecules secreted into the bloodstream. Paracrine
signaling uses molecules diffusing between two cells to communicate.
Autocrine is a cell sending a signal to itself by secreting a molecule
that binds to a receptor on its surface. Forms of communication can be
through:
Ion channels:
Can be of different types such as voltage or ligand gated ion channels.
The allow for the outflow and inflow of molecules and ions.
G-protein coupled receptor
(GPCR): Is widely recognized to contain 7 transmembrane domains. The
ligand binds on the extracellular domain and once the ligand binds, this
signals a guanine exchange factor to convert GDP to GTP and activate
the G-α subunit. G-α can target other proteins such as adenyl cyclase or
phospholipase C, which ultimately produce secondary messengers such as
cAMP, Ip3, DAG, and calcium. These secondary messengers function to
amplify signals and can target ion channels or other enzymes. One
example for amplification of a signal is cAMP binding to and activating
PKA by removing the regulatory subunits and releasing the catalytic
subunit. The catalytic subunit has a nuclear localization sequence which
prompts it to go into the nucleus and phosphorylate other proteins to
either repress or activate gene activity.
Receptor tyrosine kinases:
Bind growth factors, further promoting the tyrosine on the
intracellular portion of the protein to cross phosphorylate. The
phosphorylated tyrosine becomes a landing pad for proteins containing an
SH2 domain allowing for the activation of Ras and the involvement of
the MAP kinase pathway.
The growth process of the cell does not refer to the size of the
cell, but the density of the number of cells present in the organism at a
given time. Cell growth
pertains to the increase in the number of cells present in an organism
as it grows and develops; as the organism gets larger so does the number
of cells present. Cells are the foundation of all organisms and are the
fundamental unit of life. The growth and development of cells are
essential for the maintenance of the host and survival of the organism.
For this process, the cell goes through the steps of the cell cycle and development which involves cell growth, DNA replication, cell division, regeneration, and cell death.
The cell cycle is divided into four distinct phases: G1, S, G2, and M.
The G phase – which is the cell growth phase – makes up approximately
95% of the cycle. The proliferation of cells is instigated by
progenitors. All cells start out in an identical form and can
essentially become any type of cells. Cell signaling such as induction
can influence nearby cells to differentiate determinate the type of cell
it will become. Moreover, this allows cells of the same type to
aggregate and form tissues, then organs, and ultimately systems. The G1,
G2, and S phase (DNA replication, damage and repair) are considered to
be the interphase portion of the cycle, while the M phase (mitosis) is the cell division
portion of the cycle. Mitosis is composed of many stages which include,
prophase, metaphase, anaphase, telophase, and cytokinesis,
respectively. The ultimate result of mitosis is the formation of two
identical daughter cells.
The cell cycle is regulated by a series of signaling factors and complexes such as cyclins, cyclin-dependent kinase, and p53.
When the cell has completed its growth process and if it is found to be
damaged or altered, it undergoes cell death, either by apoptosis or necrosis, to eliminate the threat it can cause to the organism's survival.
Pathology
The scientific branch that studies and diagnoses diseases on the cellular level is called cytopathology. Cytopathology is generally used on samples of free cells or tissue fragments, in contrast to the pathology branch of histopathology,
which studies whole tissues. Cytopathology is commonly used to
investigate diseases involving a wide range of body sites, often to aid
in the diagnosis of cancer but also in the diagnosis of some infectious
diseases and other inflammatory conditions. For example, a common
application of cytopathology is the Pap smear, a screening test used to detect cervical cancer, and precancerous cervical lesions that may lead to cervical cancer.
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