Editing S phase

{{short description|DNA replication phase of the cell cycle, between G1 and G2 phase}}
{{More citations needed|date=December 2010}}

[[File:Asymmetry in the synthesis of leading and lagging strands.svg|thumb|Asymmetry in the synthesis of leading and lagging strands]]

'''S phase''' ('''Synthesis phase''') is the phase of the [[cell cycle]] in which [[DNA]] is [[DNA replication|replicated]], occurring between [[G1 phase|G<sub>1</sub> phase]] and [[G2 phase|G<sub>2</sub> phase]].<ref name=":5">{{Cite book|title=The cell cycle : principles of control|last=David |first=Morgan | name-list-style = vanc |date=2007|publisher=Oxford University Press|isbn=978-0199206100|oclc=813540567}}</ref> Since accurate duplication of the genome is critical to successful cell division, the processes that occur during S-phase are tightly regulated and widely conserved.

==Regulation==
{{Main|G1/S transition}}
Entry into S-phase is controlled by the G1 [[restriction point]] (R), which commits cells to the remainder of the cell-cycle if there is adequate nutrients and growth signaling.<ref name=":6">{{Cite book|url=https://www.ncbi.nlm.nih.gov/books/NBK6318/|title=The Restriction Point of the Cell Cycle|last1=Pardee|first1=Arthur B.|last2=Blagosklonny|first2=Mikhail V. | name-list-style = vanc |date=2013|publisher=Landes Bioscience}}</ref> This transition is essentially irreversible; after passing the restriction point, the cell will progress through S-phase even if environmental conditions become unfavorable.<ref name=":6" />

Accordingly, entry into S-phase is controlled by molecular pathways that facilitate a rapid, unidirectional shift in cell state. In yeast, for instance, cell growth induces accumulation of Cln3 [[cyclin]], which complexes with the [[Cyclin-dependent kinase|cyclin dependent kinase]] CDK2.<ref name=":7">{{cite journal | vauthors = Bertoli C, Skotheim JM, de Bruin RA | title = Control of cell cycle transcription during G1 and S phases | journal = Nature Reviews. Molecular Cell Biology | volume = 14 | issue = 8 | pages = 518–28 | date = August 2013 | pmid = 23877564 | pmc = 4569015 | doi = 10.1038/nrm3629 }}</ref> The Cln3-CDK2 complex promotes transcription of S-phase genes by inactivating the transcriptional repressor [[Whi5]].<ref name=":7" /> Since upregulation of S-phase genes drive further suppression of [[Whi5]], this pathway creates a positive feedback loop that fully commits cells to S-phase gene expression.<ref name=":7" />

A remarkably similar regulatory scheme exists in mammalian cells.<ref name=":7" /> [[Mitogen]]ic signals received throughout G1-phase cause gradual accumulation of cyclin D, which complexes with CDK4/6.<ref name=":7" /> Active cyclin D-CDK4/6 complex induces release of [[E2F]] transcription factor, which in turn initiates expression of S-phase genes.<ref name=":7" /> Several E2F target genes promote further release of E2F, creating a positive feedback loop similar to the one found in yeast.<ref name=":7" />

==DNA replication==
{{Main|DNA replication}}
[[File:Steps in DNA synthesis.svg|thumb|Steps in DNA synthesis]]

Throughout M phase and G1 phase, cells assemble inactive [[pre-replication complex]]es (pre-RC) on [[Origin of replication|replication origins]] distributed throughout the genome.<ref name=":2">{{cite journal | vauthors = Takeda DY, Dutta A | title = DNA replication and progression through S phase | journal = Oncogene | volume = 24 | issue = 17 | pages = 2827–43 | date = April 2005 | pmid = 15838518 | doi = 10.1038/sj.onc.1208616 | doi-access = free }}</ref> During S-phase, the cell converts pre-RCs into active replication forks to initiate DNA replication.<ref name=":2" /> This process depends on the kinase activity of [[Cell division cycle 7-related protein kinase|Cdc7]] and various S-phase CDKs, both of which are upregulated upon S-phase entry.<ref name=":2" />

Activation of the pre-RC is a closely regulated and highly sequential process. After Cdc7 and S-phase CDKs phosphorylate their respective substrates, a second set of replicative factors associate with the pre-RC.<ref name=":2" /> Stable association encourages [[Minichromosome maintenance|MCM helicase]] to unwind a small stretch of parental DNA into two strands of ssDNA, which in turn recruits replication protein A ([[Replication protein A|RPA]]), an ssDNA binding protein.<ref name=":2" /> RPA recruitment primes the replication fork for loading of replicative DNA [[DNA polymerase|polymerases]] and [[Proliferating cell nuclear antigen|PCNA]] sliding clamps.<ref name=":2" /> Loading of these factors completes the active replication fork and initiates synthesis of new DNA.

Complete replication fork assembly and activation only occurs on a small subset of replication origins. All eukaryotes possess many more replication origins than strictly needed during one cycle of DNA replication.<ref name=":3">{{cite journal | vauthors = Leonard AC, Méchali M | title = DNA replication origins | journal = Cold Spring Harbor Perspectives in Biology | volume = 5 | issue = 10 | pages = a010116 | date = October 2013 | pmid = 23838439 | pmc = 3783049 | doi = 10.1101/cshperspect.a010116 }}</ref> Redundant origins may increase the flexibility of DNA replication, allowing cells to control the rate of DNA synthesis and respond to replication stress.<ref name=":3" />

== Histone synthesis ==
Since new DNA must be packaged into [[nucleosome]]s to function properly, synthesis of canonical (non-variant) [[histone]] proteins occurs alongside DNA replication. During early S-phase, the cyclin E-Cdk2 complex phosphorylates [[NPAT (gene)|NPAT]], a nuclear coactivator of histone transcription.<ref name=":0">{{cite journal | vauthors = DeRan M, Pulvino M, Greene E, Su C, Zhao J | title = Transcriptional activation of histone genes requires NPAT-dependent recruitment of TRRAP-Tip60 complex to histone promoters during the G1/S phase transition | journal = Molecular and Cellular Biology | volume = 28 | issue = 1 | pages = 435–47 | date = January 2008 | pmid = 17967892 | pmc = 2223310 | doi = 10.1128/MCB.00607-07 }}</ref> NPAT is activated by phosphorylation and recruits the Tip60 chromatin remodeling complex to the promoters of histone genes.<ref name=":0" /> Tip60 activity removes inhibitory chromatin structures and drives a three to ten-fold increase in transcription rate.<ref name=":5" /><ref name=":0" />

In addition to increasing transcription of histone genes, S-phase entry also regulates histone production at the RNA level. Instead of [[Polyadenylation|polyadenylated tails]], canonical histone transcripts possess a conserved 3` [[Stem-loop|stem loop]] motif that selective binds to Stem Loop Binding Protein ([[SLBP]]).<ref name=":1">{{cite journal | vauthors = Marzluff WF, Koreski KP | title = Birth and Death of Histone mRNAs | journal = Trends in Genetics | volume = 33 | issue = 10 | pages = 745–759 | date = October 2017 | pmid = 28867047 | pmc = 5645032 | doi = 10.1016/j.tig.2017.07.014 }}</ref> SLBP binding is required for efficient processing, export, and translation of histone mRNAs, allowing it to function as a highly sensitive biochemical "switch".<ref name=":1" /> During S-phase, accumulation of SLBP acts together with NPAT to drastically increase the efficiency of histone production.<ref name=":1" /> However, once S-phase ends, both SLBP and bound RNA are rapidly degraded.<ref>{{cite journal | vauthors = Whitfield ML, Zheng LX, Baldwin A, Ohta T, Hurt MM, Marzluff WF | title = Stem-loop binding protein, the protein that binds the 3' end of histone mRNA, is cell cycle regulated by both translational and posttranslational mechanisms | journal = Molecular and Cellular Biology | volume = 20 | issue = 12 | pages = 4188–98 | date = June 2000 | pmid = 10825184 | pmc = 85788 | doi = 10.1128/MCB.20.12.4188-4198.2000 }}</ref> This immediately halts histone production and prevents a toxic buildup of free histones.<ref>{{cite journal | vauthors = Ma Y, Kanakousaki K, Buttitta L | title = How the cell cycle impacts chromatin architecture and influences cell fate | language = en | journal = Frontiers in Genetics | volume = 6 | pages = 19 | date = 2015 | pmid = 25691891 | pmc = 4315090 | doi = 10.3389/fgene.2015.00019 | doi-access = free }}</ref>

== Nucleosome replication ==
[[File:NucleosomeDuplication.png|thumb|239x239px|Conservative reassembly of core H3/H4 nucleosome behind the replication fork.|alt=]]
Free histones produced by the cell during S-phase are rapidly incorporated into new nucleosomes. This process is closely tied to the replication fork, occurring immediately in “front” and “behind” the replication complex. Translocation of MCM helicase along the leading strand disrupts parental nucleosome octamers, resulting in the release of H3-H4 and H2A-H2B subunits.<ref name=":8">{{cite journal | vauthors = Ramachandran S, Henikoff S | title = Replicating Nucleosomes | journal = Science Advances | volume = 1 | issue = 7 | pages = e1500587 | date = August 2015 | pmid = 26269799 | pmc = 4530793 | doi = 10.1126/sciadv.1500587 | bibcode = 2015SciA....1E0587R }}</ref> Reassembly of nucleosomes behind the replication fork is mediated by chromatin assembly factors (CAFs) that are loosely associated with replication proteins.<ref name=":2" /><ref name=":9">{{cite journal | vauthors = Annunziato AT | title = Split decision: what happens to nucleosomes during DNA replication? | journal = The Journal of Biological Chemistry | volume = 280 | issue = 13 | pages = 12065–8 | date = April 2005 | pmid = 15664979 | doi = 10.1074/jbc.R400039200 | doi-access = free }}</ref> Though not fully understood, the reassembly does not appear to utilize the [[Semiconservative replication|semi-conservative]] scheme seen in DNA replication.<ref name=":9" /> Labeling experiments indicate that nucleosome duplication is predominantly conservative.<ref name=":9" /><ref name=":8" /> The paternal H3-H4 core nucleosome remains completely segregated from newly synthesized H3-H4, resulting in the formation of nucleosomes that either contain exclusively old H3-H4 or exclusively new H3-H4.<ref name=":8" /><ref name=":9" /> “Old” and “new” histones are assigned to each daughter strand semi-randomly, resulting in equal division of regulatory modifications.<ref name=":8" />

== Reestablishment of chromatin domains ==
Immediately after division, each daughter chromatid only possesses half the epigenetic modifications present in the paternal chromatid.<ref name=":8" /> The cell must use this partial set of instructions to re-establish functional chromatin domains before entering mitosis.

For large genomic regions, inheritance of old H3-H4 nucleosomes is sufficient for accurate re-establishment of chromatin domains.<ref name=":8" /> Polycomb Repressive Complex 2 ([[PRC2]]) and several other histone-modifying complexes can "copy" modifications present on old histones onto new histones.<ref name=":8" /> This process amplifies epigenetic marks and counters the dilutive effect of nucleosome duplication.<ref name=":8" />

However, for small domains approaching the size of individual genes, old nucleosomes are spread too thinly for accurate propagation of histone modifications.<ref name=":8" /> In these regions, chromatin structure is probably controlled by incorporation of histone variants during nucleosome reassembly.<ref name=":8" /> The close correlation seen between H3.3/H2A.Z and transcriptionally active regions lends support to this proposed mechanism.<ref name=":8" /> Unfortunately, a causal relationship has yet to be proven.<ref name=":8" />

== DNA damage checkpoints ==
During S-phase, the cell continuously scrutinizes its genome for abnormalities. Detection of DNA damage induces activation of three canonical S-phase "checkpoint pathways" that delay or arrest further cell cycle progression:<ref name=":10">{{cite journal | vauthors = Bartek J, Lukas C, Lukas J | title = Checking on DNA damage in S phase | journal = Nature Reviews. Molecular Cell Biology | volume = 5 | issue = 10 | pages = 792–804 | date = October 2004 | pmid = 15459660 | doi = 10.1038/nrm1493 | s2cid = 33560392 }}</ref>

# The ''Replication Checkpoint'' detects stalled replication forks by integrating signals from RPA, ATR Interacting Protein (ATRIP), and RAD17.<ref name=":10" /> Upon activation, the replication checkpoint upregulates nucleotide biosynthesis and blocks replication initiation from unfired origins.<ref name=":10" /> Both of these processes contribute to rescue of stalled forks by increasing the availability of dNTPs.<ref name=":10" />
# The ''S-M Checkpoint'' blocks mitosis until the entire genome has been successfully duplicated.<ref name=":10" /> This pathway induces arrest by inhibiting the Cyclin-B-CDK1 complex, which gradually accumulates throughout the cell cycle to promote mitotic entry.<ref name=":10" />
# The ''intra-S Phase Checkpoint'' detects [[DNA repair|Double Strand Breaks]] (DSBs) through activation of [[ATR kinase|ATR]] and [[ATM kinase|ATM]] kinases.<ref name=":10" /> In addition to facilitating DNA repair, active ATR and ATM stalls cell cycle progression by promoting degradation of CDC25A, a phosphatase that removes inhibitory phosphate residues from CDKs.<ref name=":10" />  [[Homologous recombination]], an accurate process for [[DNA repair|repairing DNA]] double-strand breaks, is most active in S phase, declines in G2/M and is nearly absent in [[G1 phase]].<ref name="pmid18769152">{{cite journal |vauthors=Mao Z, Bozzella M, Seluanov A, Gorbunova V |title=DNA repair by nonhomologous end joining and homologous recombination during cell cycle in human cells |journal=Cell Cycle |volume=7 |issue=18 |pages=2902–6 |date=September 2008 |pmid=18769152 |pmc=2754209 |doi=10.4161/cc.7.18.6679 }}</ref>

In addition to these canonical checkpoints, recent evidence suggests that abnormalities in histone supply and nucleosome assembly can also alter S-phase progression.<ref name=":4">{{cite journal | vauthors = Günesdogan U, Jäckle H, Herzig A | title = Histone supply regulates S phase timing and cell cycle progression | journal = eLife | volume = 3 | pages = e02443 | date = September 2014 | pmid = 25205668 | pmc = 4157229 | doi = 10.7554/eLife.02443 | doi-access = free }}</ref> Depletion of free histones in ''Drosophila'' cells dramatically prolongs S-phase and causes permanent arrest in G2-phase.<ref name=":4" /> This unique arrest phenotype is not associated with activation of canonical DNA damage pathways, indicating that nucleosome assembly and histone supply may be scrutinized by a novel S-phase checkpoint.<ref name=":4" />

== See also ==
* [[S phase index (SPI)]]
* [[S-fraction]] or [[S-phase fraction]] (oncology/pathology prognosis)
*[[Restriction point]]

== References ==
{{reflist}}
{{Cell cycle}}

{{DEFAULTSORT:S Phase}}
[[Category:Cell cycle]]