The G
0 phase describes a cellular state outside of the replicative
cell cycle
The cell cycle, or cell-division cycle, is the series of events that take place in a cell that cause it to divide into two daughter cells. These events include the duplication of its DNA (DNA replication) and some of its organelles, and subs ...
. Classically, cells were thought to enter G
0 primarily due to environmental factors, like nutrient deprivation, that limited the resources necessary for proliferation. Thus it was thought of as a ''resting phase''. G
0 is now known to take different forms and occur for multiple reasons. For example, most adult
neuronal cells, among the most metabolically active cells in the body, are fully differentiated and reside in a terminal G
0 phase. Neurons reside in this state, not because of stochastic or limited nutrient supply, but as a part of their developmental program.
G
0 was first suggested as a cell state based on early cell cycle studies. When the first studies defined the four phases of the cell cycle using radioactive labeling techniques, it was discovered that not all cells in a population
proliferate at similar rates.
A population's “growth fraction” – or the fraction of the population that was growing – was actively proliferating, but other cells existed in a non-proliferative state. Some of these non-proliferating cells could respond to extrinsic stimuli and proliferate by re-entering the cell cycle.
Early contrasting views either considered non-proliferating cells to simply be in an extended
G1 phase or in a cell cycle phase distinct from G
1 – termed G
0.
Subsequent research pointed to a
restriction point (R-point) in G
1 where cells can enter G
0 before the R-point but are committed to mitosis after the R-point.
These early studies provided evidence for the existence of a G
0 state to which access is restricted. These cells that do not divide further exit G1 phase to enter an inactive stage called quiescent stage.
Diversity of G0 states
Three G
0 states exist and can be categorized as either reversible (
quiescent
Quiescence (/kwiˈɛsəns/) is a state of quietness or inactivity. It may refer to:
* Quiescence search, in game tree searching (adversarial search) in artificial intelligence, a quiescent state is one in which a game is considered stable and unl ...
) or irreversible (
senescent and
differentiated). Each of these three states can be entered from the G
1 phase before the cell commits to the next round of the cell cycle. Quiescence refers to a reversible G
0 state where subpopulations of cells reside in a 'quiescent' state before entering the cell cycle after activation in response to extrinsic signals. Quiescent cells are often identified by low
RNA content, lack of cell proliferation markers, and increased label retention indicating low cell turnover.
Senescence is distinct from quiescence because senescence is an irreversible state that cells enter in response to DNA damage or degradation that would make a cell's progeny nonviable. Such
DNA damage can occur from
telomere shortening over many cell divisions as well as reactive oxygen species (ROS) exposure, oncogene activation, and cell-cell fusion. While senescent cells can no longer replicate, they remain able to perform many normal cellular functions. Senescence is often a biochemical alternative to the self-destruction of such a damaged cell by
apoptosis
Apoptosis (from grc, ἀπόπτωσις, apóptōsis, 'falling off') is a form of programmed cell death that occurs in multicellular organisms. Biochemical events lead to characteristic cell changes (morphology) and death. These changes includ ...
. In contrast to cellular senescence, quiescence is not a reactive event but part of the core programming of several different cell types. Finally, differentiated cells are stem cells that have progressed through a differentiation program to reach a mature – terminally differentiated – state. Differentiated cells continue to stay in G
0 and perform their main functions indefinitely.
Characteristics of quiescent stem cells
Transcriptomes
The
transcriptomes of several types of quiescent stem cells, such as
hematopoietic
Haematopoiesis (, from Greek , 'blood' and 'to make'; also hematopoiesis in American English; sometimes also h(a)emopoiesis) is the formation of blood cellular components. All cellular blood components are derived from haematopoietic stem cells ...
, muscle, and hair follicle, have been characterized through
high-throughput techniques, such as
microarray and
RNA sequencing
RNA-Seq (named as an abbreviation of RNA sequencing) is a sequencing technique which uses next-generation sequencing (NGS) to reveal the presence and quantity of RNA in a biological sample at a given moment, analyzing the continuously changing c ...
. Although variations exist in their individual transcriptomes, most quiescent tissue stem cells share a common pattern of gene expression that involves downregulation of cell cycle progression genes, such as
cyclin A2
Cyclin-A2 is a protein that in humans is encoded by the ''CCNA2'' gene. It is one of the two types of cyclin A: cyclin A1 is expressed during meiosis and embryogenesis while cyclin A2 is expressed in dividing somatic cells.
Function
Cyclin A2 ...
,
cyclin B1,
cyclin E2
Cyclin E2 is a protein that in humans is encoded by the ''CCNE2'' gene. It is a G1 cyclin that binds Cdk2 and is inhibited by p27(Kip1) and p21(Cip1). It plays a role in the G1/S portion of the cell cycle and also has putative interactions wit ...
, and
survivin, and upregulation of genes involved in the regulation of transcription and stem cell fate, such as
FOXO3
Forkhead box O3, also known as FOXO3 or FOXO3a, is a human protein encoded by the ''FOXO3'' gene.
Function
FOXO3 belongs to the O subclass of the forkhead family of transcription factors which are characterized by a distinct fork head DNA-b ...
and
EZH1 #REDIRECT EZH1
{{R from other capitalization
...
. Downregulation of mitochondrial
cytochrome C
The cytochrome complex, or cyt ''c'', is a small hemeprotein found loosely associated with the inner membrane of the mitochondrion. It belongs to the cytochrome c family of proteins and plays a major role in cell apoptosis. Cytochrome c is high ...
also reflects the low metabolic state of quiescent stem cells.
Epigenetic
Many quiescent stem cells, particularly
adult stem cells
Adult stem cells are undifferentiated cells, found throughout the body after development, that multiply by cell division to replenish dying cells and regenerate damaged tissues. Also known as somatic stem cells (from Greek σωματικóς, ...
, also share similar
epigenetic
In biology, epigenetics is the study of stable phenotypic changes (known as ''marks'') that do not involve alterations in the DNA sequence. The Greek prefix '' epi-'' ( "over, outside of, around") in ''epigenetics'' implies features that are "o ...
patterns. For example,
H3K4me3 and
H3K27me3
H3K27me3 is an epigenetic modification to the DNA packaging protein Histone H3. It is a mark that indicates the tri-methylation of lysine 27 on histone H3 protein.
This tri-methylation is associated with the downregulation of nearby genes via t ...
, are two major
histone methylation patterns that form a bivalent domain and are located near transcription initiation sites. These epigenetic markers have been found to regulate lineage decisions in embryonic stem cells as well as control quiescence in hair follicle and muscle stem cells via
chromatin
Chromatin is a complex of DNA and protein found in eukaryotic cells. The primary function is to package long DNA molecules into more compact, denser structures. This prevents the strands from becoming tangled and also plays important roles in ...
modification.
Regulation of quiescence
Cell cycle regulators
Functional
tumor suppressor genes
A tumor suppressor gene (TSG), or anti-oncogene, is a gene that regulates a cell during cell division and replication. If the cell grows uncontrollably, it will result in cancer. When a tumor suppressor gene is mutated, it results in a loss or red ...
, particularly
p53
p53, also known as Tumor protein P53, cellular tumor antigen p53 (UniProt name), or transformation-related protein 53 (TRP53) is a regulatory protein that is often mutated in human cancers. The p53 proteins (originally thought to be, and often ...
and
Rb gene, are required to maintain stem cell quiescence and prevent exhaustion of the
progenitor cell pool through excessive divisions. For example, deletion of all three components of the Rb family of proteins has been shown to halt quiescence in hematopoietic stem cells. Lack of p53 has been shown to prevent differentiation of these stem cells due to the cells’ inability to exit the cell cycle into the G
0 phase. In addition to p53 and Rb,
cyclin dependent kinase
Cyclin-dependent kinases (CDKs) are the families of protein kinases first discovered for their role in regulating the cell cycle. They are also involved in regulating transcription, mRNA processing, and the differentiation of nerve cells. They a ...
inhibitors (CKIs), such as
p21,
p27, and
p57, are also important for maintaining quiescence. In mouse hematopoietic stem cells, knockout of p57 and p27 leads to G
0 exit through nuclear import of
cyclin D1
Cyclin D1 is a protein that in humans is encoded by the ''CCND1'' gene.
Gene expression
The CCND1 gene encodes the cyclin D1 protein. The human CCND1 gene is located on the long arm of chromosome 11 (band 11q13). It is 13,388 base pairs lo ...
and subsequent
phosphorylation of Rb. Finally, the
Notch signaling pathway has been shown to play an important role in maintenance of quiescence.
Post-transcriptional regulation
Post-transcriptional regulation of gene expression via
miRNA
MicroRNA (miRNA) are small, single-stranded, non-coding RNA molecules containing 21 to 23 nucleotides. Found in plants, animals and some viruses, miRNAs are involved in RNA silencing and post-transcriptional regulation of gene expression. m ...
synthesis has been shown to play an equally important role in the maintenance of stem cell quiescence. miRNA strands bind to the 3’ untranslated region (
3’ UTR) of target
mRNA’s, preventing their translation into functional proteins. The length of the 3’ UTR of a gene determines its ability to bind to miRNA strands, thereby allowing regulation of quiescence. Some examples of miRNA's in stem cells include miR-126, which controls the
PI3K/AKT/mTOR pathway
The PI3K/AKT/mTOR pathway is an intracellular signaling pathway important in regulating the cell cycle. Therefore, it is directly related to cellular quiescence, proliferation, cancer, and longevity. PI3K activation phosphorylates and activates ...
in hematopoietic stem cells, miR-489, which suppresses the DEK
oncogene in muscle stem cells, and miR-31, which regulates
Myf5 in muscle stem cells. miRNA sequestration of mRNA within
ribonucleoprotein
Nucleoproteins are proteins conjugated with nucleic acids (either DNA or RNA). Typical nucleoproteins include ribosomes, nucleosomes and viral nucleocapsid proteins.
Structures
Nucleoproteins tend to be positively charged, facilitating int ...
complexes allows quiescent cells to store the mRNA necessary for quick entry into the
G1 phase.
Response to stress
Stem cells that have been quiescent for a long time often face various environmental stressors, such as
oxidative stress
Oxidative stress reflects an imbalance between the systemic manifestation of reactive oxygen species and a biological system's ability to readily detoxify the reactive intermediates or to repair the resulting damage. Disturbances in the normal r ...
. However, several mechanisms allow these cells to respond to such stressors. For example, the
FOXO
FOX (forkhead box) proteins are a family of transcription factors that play important roles in regulating the expression of genes involved in cell growth, proliferation, differentiation, and longevity. Many FOX proteins are important to embryonic ...
transcription factors respond to the presence of
reactive oxygen species (ROS) while
HIF1A
Hypoxia-inducible factor 1-alpha, also known as HIF-1-alpha, is a subunit of a heterodimeric transcription factor hypoxia-inducible factor 1 ( HIF-1) that is encoded by the ''HIF1A'' gene. The Nobel Prize in Physiology or Medicine 2019 was awarde ...
and
LKB1 respond to
hypoxic conditions. In hematopoietic stem cells,
autophagy
Autophagy (or autophagocytosis; from the Ancient Greek , , meaning "self-devouring" and , , meaning "hollow") is the natural, conserved degradation of the cell that removes unnecessary or dysfunctional components through a lysosome-dependent r ...
is induced to respond to metabolic stress.
Examples of reversible G0 phase
Tissue stem cells
Stem cells are cells with the unique ability to produce differentiated
daughter cells and to preserve their stem cell identity through self-renewal.
In mammals, most adult tissues contain
tissue-specific stem cells that reside in the tissue and proliferate to maintain homeostasis for the lifespan of the organism. These cells can undergo immense proliferation in response to tissue damage before differentiating and engaging in regeneration. Some tissue stem cells exist in a reversible, quiescent state indefinitely until being activated by external stimuli. Many different types of tissue stem cells exist, including
muscle stem cells (MuSCs),
neural stem cells (NSCs),
intestinal stem cells (ISCs), and many others.
Stem cell quiescence has been recently suggested to be composed of two distinct functional phases, G
0 and an ‘alert’ phase termed G
Alert.
Stem cells are believed to actively and reversibly transition between these phases to respond to injury stimuli and seem to gain enhanced tissue regenerative function in G
Alert. Thus, transition into G
Alert has been proposed as an adaptive response that enables stem cells to rapidly respond to injury or stress by priming them for cell cycle entry. In muscle stem cells,
mTORC1 activity has been identified to control the transition from G
0 into G
Alert along with signaling through the
HGF receptor
cMet.
Mature hepatocytes
While a reversible quiescent state is perhaps most important for tissue stem cells to respond quickly to stimuli and maintain proper homeostasis and regeneration, reversible G
0 phases can be found in non-stem cells such as mature hepatocytes.
Hepatocytes are typically quiescent in normal livers but undergo limited replication (less than 2 cell divisions) during liver regeneration after partial hepatectomy. However, in certain cases, hepatocytes can experience immense proliferation (more than 70 cell divisions) indicating that their proliferation capacity is not hampered by existing in a reversible quiescent state.
Examples of irreversible G0 phase
Senescent cells
Often associated with aging and age-related diseases in vivo, senescent cells can be found in many renewable tissues, including the
stroma,
vasculature,
hematopoietic system, and many
epithelial
Epithelium or epithelial tissue is one of the four basic types of animal tissue, along with connective tissue, muscle tissue and nervous tissue. It is a thin, continuous, protective layer of compactly packed cells with a little intercellula ...
organs. Resulting from accumulation over many cell divisions, senescence is often seen in age-associated degenerative phenotypes. Senescent fibroblasts in models of breast epithelial cell function have been found to disrupt milk protein production due to secretion of matrix
metalloproteinases.
Similarly, senescent pulmonary artery smooth muscle cells caused nearby smooth muscle cells to proliferate and migrate, perhaps contributing to hypertrophy of pulmonary arteries and eventually pulmonary hypertension.
Differentiated muscle
During skeletal
myogenesis
Myogenesis is the formation of skeletal muscular tissue, particularly during embryonic development.
Muscle fibers generally form through the fusion of precursor myoblasts into multinucleated fibers called ''myotubes''. In the early development ...
, cycling progenitor cells known as
myoblast
Myogenesis is the formation of skeletal muscular tissue, particularly during embryonic development.
Muscle fibers generally form through the fusion of precursor myoblasts into multinucleated fibers called ''myotubes''. In the early development of ...
s differentiate and fuse together into non-cycling muscle cells called myocytes that remain in a terminal G
0 phase.
[page 395, Biology, Fifth Edition, Campbell, 1999] As a result, the fibers that make up
skeletal muscle (myofibers) are cells with multiple nuclei, referred to as myonuclei, since each myonucleus originated from a single myoblast. Skeletal muscle cells continue indefinitely to provide contractile force through simultaneous contractions of cellular structures called
sarcomeres. Importantly, these cells are kept in a terminal G
0 phase since disruption of muscle fiber structure after myofiber formation would prevent proper transmission of force through the length of the muscle. Muscle growth can be stimulated by growth or injury and involves the recruitment of muscle stem cells – also known as satellite cells – out of a reversible quiescent state. These stem cells differentiate and fuse to generate new muscle fibers both in parallel and in series to increase force generation capacity.
Cardiac muscle
Cardiac muscle (also called heart muscle, myocardium, cardiomyocytes and cardiac myocytes) is one of three types of vertebrate muscle tissues, with the other two being skeletal muscle and smooth muscle. It is an involuntary, striated muscle that ...
is also formed through myogenesis but instead of recruiting stem cells to fuse and form new cells, heart muscle cells – known as
cardiomyocyte
Cardiac muscle (also called heart muscle, myocardium, cardiomyocytes and cardiac myocytes) is one of three types of vertebrate muscle tissues, with the other two being skeletal muscle and smooth muscle. It is an involuntary, striated muscle that ...
s – simply increase in size as the heart grows larger. Similarly to skeletal muscle, if cardiomyocytes had to continue dividing to add muscle tissue the contractile structures necessary for heart function would be disrupted.
Differentiated bone
Of the four major types of bone cells,
osteocyte
An osteocyte, an oblate shaped type of bone cell with dendritic processes, is the most commonly found cell in mature bone. It can live as long as the organism itself. The adult human body has about 42 billion of them. Osteocytes do not divide and ...
s are the most common and also exist in a terminal G
0 phase. Osteocytes arise from osteoblasts that are trapped within a self-secreted matrix. While osteocytes also have reduced synthetic activity, they still serve bone functions besides generating structure. Osteocytes work through various mechanosensory mechanisms to assist in the routine turnover over bony matrix.
Differentiated nerve
Outside of a few
neurogenic
In biology, the nervous system is the highly complex part of an animal that coordinates its actions and sensory information by transmitting signals to and from different parts of its body. The nervous system detects environmental changes ...
niches in the brain, most
neurons are fully differentiated and reside in a terminal G
0 phase. These fully differentiated neurons form
synapses where electrical signals are transmitted by
axon
An axon (from Greek ἄξων ''áxōn'', axis), or nerve fiber (or nerve fibre: see spelling differences), is a long, slender projection of a nerve cell, or neuron, in vertebrates, that typically conducts electrical impulses known as action p ...
s to the
dendrite
Dendrites (from Greek δένδρον ''déndron'', "tree"), also dendrons, are branched protoplasmic extensions of a nerve cell that propagate the electrochemical stimulation received from other neural cells to the cell body, or soma, of the n ...
s of nearby neurons. In this G
0 state, neurons continue functioning until senescence or apoptosis. Numerous studies have reported accumulation of
DNA damage with age, particularly
oxidative damage, in the mammalian
brain.
[Bernstein H, Payne CM, Bernstein C, Garewal H, Dvorak K (2008). Cancer and aging as consequences of un-repaired DNA damage. In: New Research on DNA Damages (Editors: Honoka Kimura and Aoi Suzuki) ]Nova Science Publishers, Inc.
Nova Science Publishers is an academic publisher of books, encyclopedias, handbooks, e-books and journals, based in Hauppauge, New York. It was founded in 1985. A prolific publisher of books, Nova has received criticism from librarians for not a ...
, New York, Chapter 1, pp. 1–47. open access, but read only https://www.novapublishers.com/catalog/product_info.php?products_id=43247
Mechanism of G0 entry
Role of Rim15
Rim15 was first discovered to play a critical role in initiating
meiosis in
diploid yeast cells. Under conditions of low glucose and nitrogen, which are key nutrients for the survival of yeast, diploid yeast cells initiate meiosis through the activation of early meiotic-specific genes (EMGs). The expression of EMGs is regulated by Ume6. Ume6 recruits the
histone deacetylases,
Rpd3 and Sin3, to repress EMG expression when glucose and nitrogen levels are high, and it recruits the EMG transcription factor Ime1 when glucose and nitrogen levels are low. Rim15, named for its role in the regulation of an EMG called IME2, displaces Rpd3 and Sin3, thereby allowing Ume6 to bring Ime1 to the promoters of EMGs for meiosis initiation.
In addition to playing a role in meiosis initiation, Rim15 has also been shown to be a critical effector for yeast cell entry into G
0 in the presence of stress. Signals from several different nutrient signaling pathways converge on Rim15, which activates the transcription factors, Gis1, Msn2, and Msn4. Gis1 binds to and activates promoters containing post-
diauxic growth shift (PDS) elements while Msn2 and Msn4 bind to and activate promoters containing stress-
response elements Response elements are short sequences of DNA within a gene promoter or enhancer region that are able to bind specific transcription factors and regulate transcription of genes.
Under conditions of stress, a transcription activator protein binds ...
(STREs). Although it is not clear how Rim15 activates Gis1 and Msn2/4, there is some speculation that it may directly phosphorylate them or be involved in chromatin remodeling. Rim15 has also been found to contain a
PAS domain
A Per-Arnt-Sim (PAS) domain is a protein domain found in all kingdoms of life. Generally, the PAS domain acts as a molecular sensor, whereby small molecules and other proteins associate via binding of the PAS domain. Due to this sensing capability ...
at its
N terminal, making it a newly discovered member of the PAS
kinase family. The PAS domain is a regulatory unit of the Rim15 protein that may play a role in sensing oxidative stress in yeast.
Nutrient signaling pathways
Glucose
Yeast grows exponentially through
fermentation of glucose. When glucose levels drop, yeast shift from fermentation to
cellular respiration
Cellular respiration is the process by which biological fuels are oxidised in the presence of an inorganic electron acceptor such as oxygen to produce large amounts of energy, to drive the bulk production of ATP. Cellular respiration may be des ...
, metabolizing the fermentative products from their exponential growth phase. This shift is known as the diauxic shift after which yeast enter G
0. When glucose levels in the surroundings are high, the production of
cAMP
Camp may refer to:
Outdoor accommodation and recreation
* Campsite or campground, a recreational outdoor sleeping and eating site
* a temporary settlement for nomads
* Camp, a term used in New England, Northern Ontario and New Brunswick to descri ...
through the RAS-cAMP-PKA pathway (a
cAMP-dependent pathway
In the field of molecular biology, the cAMP-dependent pathway, also known as the adenylyl cyclase pathway, is a G protein-coupled receptor-triggered signaling cascade used in cell communication.
Discovery
cAMP was discovered by Earl Sutherlan ...
) is elevated, causing
protein kinase A
In cell biology, protein kinase A (PKA) is a family of enzymes whose activity is dependent on cellular levels of cyclic AMP (cAMP). PKA is also known as cAMP-dependent protein kinase (). PKA has several functions in the cell, including regulatio ...
(PKA) to inhibit its downstream target Rim15 and allow cell proliferation. When glucose levels drop, cAMP production declines, lifting PKA's inhibition of Rim15 and allowing the yeast cell to enter G
0.
Nitrogen
In addition to glucose, the presence of nitrogen is crucial for yeast proliferation. Under low nitrogen conditions, Rim15 is activated to promote cell cycle arrest through inactivation of the protein kinases
TORC1 and Sch9. While TORC1 and Sch9 belong to two separate pathways, namely the TOR and Fermentable Growth Medium induced pathways respectively, both protein kinases act to promote cytoplasmic retention of Rim15. Under normal conditions, Rim15 is anchored to the cytoplasmic
14-3-3 protein
14-3-3 proteins are a family of conserved regulatory molecules that are expressed in all eukaryotic cells. 14-3-3 proteins have the ability to bind a multitude of functionally diverse signaling proteins, including kinases, phosphatases, and tran ...
, Bmh2, via phosphorylation of its Thr1075. TORC1 inactivates certain
phosphatases
In biochemistry, a phosphatase is an enzyme that uses water to cleave a phosphoric acid monoester into a phosphate ion and an alcohol. Because a phosphatase enzyme catalyzes the hydrolysis of its substrate, it is a subcategory of hydrolases. P ...
in the cytoplasm, keeping Rim15 anchored to Bmh2, while it is thought that Sch9 promotes Rim15 cytoplasmic retention through phosphorylation of another 14-3-3 binding site close to Thr1075. When extracellular nitrogen is low, TORC1 and Sch9 are inactivated, allowing dephosphorylation of Rim15 and its subsequent transport to the nucleus, where it can activate transcription factors involved in promoting cell entry into G
0. It has also been found that Rim15 promotes its own export from the nucleus through
autophosphorylation.
Phosphate
Yeast cells respond to low extracellular phosphate levels by activating genes that are involved in the production and upregulation of inorganic phosphate. The PHO pathway is involved in the regulation of phosphate levels. Under normal conditions, the yeast
cyclin-dependent kinase complex, Pho80-Pho85, inactivates the
Pho4 transcription factor through phosphorylation. However, when phosphate levels drop, Pho81 inhibits Pho80-Pho85, allowing Pho4 to be active. When phosphate is abundant, Pho80-Pho85 also inhibits the nuclear pool of Rim 15 by promoting phosphorylation of its Thr1075 Bmh2 binding site. Thus, Pho80-Pho85 acts in concert with Sch9 and TORC1 to promote cytoplasmic retention of Rim15 under normal conditions.
Mechanism of G0 exit
Cyclin C/Cdk3 and Rb
The transition from G
1 to
S phase is promoted by the inactivation of Rb through its progressive
hyperphosphorylation by the
Cyclin D/Cdk4 and
Cyclin E/Cdk2 complexes in late G
1. An early observation that loss of Rb promoted cell cycle re-entry in G
0 cells suggested that Rb is also essential in regulating the G
0 to G
1 transition in quiescent cells.
Further observations revealed that levels of
cyclin C
Cyclin-C is a protein that in humans is encoded by the ''CCNC'' gene.
The protein encoded by this gene is a member of the cyclin family of proteins. The encoded protein interacts with cyclin-dependent kinase 8 and induces the phosphorylation of t ...
mRNA are highest when human cells exit G
0, suggesting that cyclin C may be involved in Rb phosphorylation to promote cell cycle re-entry of G
0 arrested cells.
Immunoprecipitation kinase assays revealed that cyclin C has Rb kinase activity. Furthermore, unlike cyclins D and E, cyclin C's Rb kinase activity is highest during early G
1 and lowest during late G
1 and S phases, suggesting that it may be involved in the G
0 to G
1 transition. The use of
fluorescence-activated cell sorting to identify G
0 cells, which are characterized by a high DNA to RNA ratio relative to G
1 cells, confirmed the suspicion that cyclin C promotes G
0 exit as repression of endogenous cyclin C by
RNAi
RNA interference (RNAi) is a biological process in which RNA molecules are involved in sequence-specific suppression of gene expression by double-stranded RNA, through translational or transcriptional repression. Historically, RNAi was known by o ...
in mammalian cells increased the proportion of cells arrested in G
0. Further experiments involving mutation of Rb at specific phosphorylation sites showed that cyclin C phosphorylation of Rb at S807/811 is necessary for G
0 exit. It remains unclear, however, whether this phosphorylation pattern is sufficient for G0 exit. Finally, co-immunoprecipitation assays revealed that
cyclin-dependent kinase 3 (cdk3) promotes G
0 exit by forming a complex with cyclin C to phosphorylate Rb at S807/811. Interestingly, S807/811 are also targets of cyclin D/cdk4 phosphorylation during the G
1 to S transition. This might suggest a possible compensation of cdk3 activity by cdk4, especially in light of the observation that G
0 exit is only delayed, and not permanently inhibited, in cells lacking cdk3 but functional in cdk4. Despite the overlap of phosphorylation targets, it seems that cdk3 is still necessary for the most effective transition from G
0 to G
1.
Rb and G0 exit
Studies suggest that Rb repression of the
E2F
E2F is a group of genes that encodes a family of transcription factors (TF) in higher eukaryotes. Three of them are activators: E2F1, 2 and E2F3a. Six others act as suppressors: E2F3b, E2F4-8. All of them are involved in the cell cycle regulation a ...
family of transcription factors regulates the G
0 to G
1 transition just as it does the G
1 to S transition. Activating E2F complexes are associated with the recruitment of
histone acetyltransferases
Histone acetyltransferases (HATs) are enzymes that acetylate conserved lysine amino acids on histone proteins by transferring an acetyl group from acetyl-CoA to form ε-''N''-acetyllysine. DNA is wrapped around histones, and, by transferring an ...
, which activate gene expression necessary for G
1 entry, while
E2F4
Transcription factor E2F4 is a protein that in humans is encoded by the ''E2F4'' gene.
Function
The protein encoded by this gene is a member of the E2F family of transcription factors. The E2F family plays a crucial role in the control of cel ...
complexes recruit histone deacetylases, which repress gene expression. Phosphorylation of Rb by Cdk complexes allows its dissociation from E2F transcription factors and the subsequent expression of genes necessary for G
0 exit. Other members of the Rb
pocket protein family
Pocket protein family consists of three proteins:
* RB – Retinoblastoma protein
* p107 – Retinoblastoma-like protein 1
* p130 – Retinoblastoma-like protein 2
They play crucial roles in the metazoan cell cycle through interaction with member ...
, such as p107 and p130, have also been found to be involved in G
0 arrest. p130 levels are elevated in G
0 and have been found to associate with E2F-4 complexes to repress transcription of E2F target genes. Meanwhile, p107 has been found to rescue the cell arrest phenotype after loss of Rb even though p107 is expressed at comparatively low levels in G
0 cells. Taken together, these findings suggest that Rb repression of E2F transcription factors promotes cell arrest while phosphorylation of Rb leads to G
0 exit via derepression of E2F target genes.
In addition to its regulation of E2F, Rb has also been shown to suppress
RNA polymerase I and
RNA polymerase III, which are involved in
rRNA
Ribosomal ribonucleic acid (rRNA) is a type of non-coding RNA which is the primary component of ribosomes, essential to all cells. rRNA is a ribozyme which carries out protein synthesis in ribosomes. Ribosomal RNA is transcribed from ribosomal ...
synthesis. Thus, phosphorylation of Rb also allows activation of rRNA synthesis, which is crucial for protein synthesis upon entry into G
1.
References
{{DEFAULTSORT:G0 Phase
Cell cycle