Cell cycle: To differentiate or not to differentiate?

Developmental regulation of the cell cycle is an important determinant of tissue size and shape. Equally important is regulated withdrawal from the cell cycle to allow cells to differentiate. Recent evidence supports a direct link between transcriptional regulation of the cell cycle machinery and cell differentiation

The Contribution of E2F-Regulated Transcription to Drosophila PCNA Gene Function

E2F proteins control cell cycle progression by predominantly acting as either activators or repressors of transcription [1]. How the antagonizing activities of different E2Fs are integrated by cis-acting control regions into a final transcriptional output in an intact animal is not well understood. E2F function is required for normal development in many species [2–7], but it is not completely clear for which genes E2F-regulated transcription provides an essential biological function. To address these questions, we have characterized the control region of the Drosophila PCNA gene. A single E2F binding site within a 100-bp enhancer is necessary and sufficient to direct the correct spatiotemporal program of G1-S-regulated PCNA expression during development. This dynamic program requires both E2F-mediated transcriptional activation and repression, which, in Drosophila, are thought to be carried out by two distinct E2F proteins [2, 3, 8–11]. Our data suggest that functional antagonism between these different E2F proteins can occur in vivo by competition for the same binding site. An engineered PCNA gene with mutated E2F binding sites supports a low level of expression that can partially rescue the lethality of PCNA null mutants. Thus, E2F regulation of PCNA is dispensable for viability, but is nonetheless important for normal Drosophila development

Coordinating cell cycle-regulated histone gene expression through assembly and function of the Histone Locus Body

Metazoan replication-dependent (RD) histone genes encode the only known cellular mRNAs that are not polyadenylated. These mRNAs end instead in a conserved stem-loop, which is formed by an endonucleolytic cleavage of the pre-mRNA. The genes for all 5 histone proteins are clustered in all metazoans and coordinately regulated with high levels of expression during S phase. Production of histone mRNAs occurs in a nuclear body called the Histone Locus Body (HLB), a subdomain of the nucleus defined by a concentration of factors necessary for histone gene transcription and pre-mRNA processing. These factors include the scaffolding protein NPAT, essential for histone gene transcription, and FLASH and U7 snRNP, both essential for histone pre-mRNA processing. Histone gene expression is activated by Cyclin E/Cdk2-mediated phosphorylation of NPAT at the G1-S transition. The concentration of factors within the HLB couples transcription with pre-mRNA processing, enhancing the efficiency of histone mRNA biosynthesis

Drosophila Roc1a Encodes a RING-H2 Protein with a Unique Function in Processing the Hh Signal Transducer Ci by the SCF E3 Ubiquitin Ligase

Substrate specificity of SCF E3 ubiquitin ligases is thought to be determined by the F box protein subunit. Another component of SCF complexes is provided by members of the Roc1/Rbx1/Hrt1 gene family, which encode RING-H2 proteins. Drosophila contains three members of this gene family. We show that Roc1a mutant cells fail to proliferate. Further, while the F box protein Slimb is required for Cubitus interruptus (Ci) and Armadillo/beta-catenin (Arm) proteolysis, Roc1a mutant cells hyperaccumulate Ci but not Arm. This suggests that Slimb and Roc1a function in the same SCF complex to target Ci but that a different RING-H2 protein acts with Slimb to target Arm. Consequently, the identity of the Roc subunit may contribute to the selection of substrates by metazoan SCF complexes

stringcdc25 and cyclin E are required for patterned histone expression at different stages of Drosophila embryonic development

Metazoan replication-dependent histone mRNAs accumulate to high levels during S phase as a result of an increase in the rate of histone gene transcription, pre-mRNA processing, and mRNA stability at the G1–S transition. However, relatively little is known about the contribution of these processes to histone expression in the cell cycles of early development, which often lack a G1 phase. In post-blastoderm Drosophila embryos, zygotic expression of the stgcdc25 phosphatase in G2 activates cyclin/cdc2 kinases and triggers mitosis. Here we show that histone transcription initiates in late G2 of cycle 14 in response to stgcdc25 and in anticipation of S phase of the next cycle, which occurs immediately following mitosis. Mutation of stgcdc25 arrests cells in G2 and prevents histone transcription. Expression of a mutant form of Cdc2 that bypasses the requirement for stgcdc25 activates histone transcription during G2 in stgcdc25 mutant embryos. Thus, in these embryonic cycles, histone transcription is controlled by the principal G2–M regulators, stringcdc25, and cdc2 kinase, rather than solely by regulators of the G1–S transition. After the introduction of G1–S control midway through embryogenesis, histone expression depends on DNA replication and the function of cyclin E, and no longer requires stgcdc25. Thus, during the altered cell cycles of early animal development, different cell cycle mechanisms are employed to ensure that the production of histones accompanies DNA synthesis

Signaling Pathways that Control Cell Proliferation

Cells decide to proliferate or remain quiescent using signaling pathways that link information about the cellular environment to the G1 phase of the cell cycle. Progression through G1 phase is controlled by pRB proteins, which function to repress the activity of E2F transcription factors in cells exiting mitosis and in quiescent cells. Phosphorylation of pRB proteins by the G1 cyclin-dependent kinases (CDKs) releases E2F factors, promoting the transition to S phase. CDK activity is primarily regulated by the binding of CDK catalytic subunits to cyclin partners and CDK inhibitors. Consequently, both mitogenic and antiproliferative signals exert their effects on cell proliferation through the transcriptional regulation and ubiquitin-dependent degradation of cyclins and CDK inhibitors

A population of G2-arrested cells are selected as sensory organ precursors for the interommatidial bristles of the Drosophila eye

Cell cycle progression and differentiation are highly coordinated during the development of multicellular organisms. The mechanisms by which these processes are coordinated and how their coordination contributes to normal development are not fully understood. Here, we determine the developmental fate of a population of precursor cells in the developing Drosophila melanogaster retina that arrest in G2 phase of the cell cycle and investigate whether cell cycle phase-specific arrest influences the fate of these cells. We demonstrate that retinal precursor cells that arrest in G2 during larval development are selected as sensory organ precursors (SOPs) during pupal development and undergo two cell divisions to generate the four-cell interommatidial mechanosensory bristles. While G2 arrest is not required for bristle development, preventing G2 arrest results in incorrect bristle positioning in the adult eye. We conclude that G2-arrested cells provide a positional cue during development to ensure proper spacing of bristles in the eye. Our results suggest that the control of cell cycle progression refines cell fate decisions and that the relationship between these two processes is not necessarily deterministic

Methylation of histone H4 lysine 20 by PR-Set7 ensures the integrity of late replicating sequence domains in Drosophila

The methylation state of lysine 20 on histone H4 (H4K20) has been linked to chromatin compaction, transcription, DNA repair and DNA replication. Monomethylation of H4K20 (H4K20me1) is mediated by the cell cycle-regulated histone methyltransferase PR-Set7. PR-Set7 depletion in mammalian cells results in defective S phase progression and the accumulation of DNA damage, which has been partially attributed to defects in origin selection and activation. However, these studies were limited to only a handful of mammalian origins, and it remains unclear how PR-Set7 and H4K20 methylation impact the replication program on a genomic scale. We employed genetic, cytological, and genomic approaches to better understand the role of PR-Set7 and H4K20 methylation in regulating DNA replication and genome stability in Drosophila cells. We find that deregulation of H4K20 methylation had no impact on origin activation throughout the genome. Instead, depletion of PR-Set7 and loss of H4K20me1 results in the accumulation of DNA damage and an ATR-dependent cell cycle arrest. Coincident with the ATR-dependent cell cycle arrest, we find increased DNA damage that is specifically limited to late replicating regions of the Drosophila genome, suggesting that PR-Set7-mediated monomethylation of H4K20 is critical for maintaining the genomic integrity of late replicating domains

Scalloped and Yorkie are required for cell cycle re-entry of quiescent cells after tissue damage

Regeneration of damaged tissues typically requires a population of active stem cells. How damaged tissue is regenerated in quiescent tissues lacking a stem cell population is less well understood. We used a genetic screen in the developing Drosophila melanogaster eye to investigate the mechanisms that trigger quiescent cells to re-enter the cell cycle and proliferate in response to tissue damage. We discovered that Hippo signaling regulates compensatory proliferation after extensive cell death in the developing eye. Scalloped and Yorkie, transcriptional effectors of the Hippo pathway, drive Cyclin E expression to induce cell cycle re-entry in cells that normally remain quiescent in the absence of damage. Ajuba, an upstream regulator of Hippo signaling that functions as a sensor of epithelial integrity, is also required for cell cycle re-entry. Thus, in addition to its well-established role in modulating proliferation during periods of tissue growth, Hippo signaling maintains homeostasis by regulating quiescent cell populations affected by tissue damage

Endoreplication and polyploidy: insights into development and disease

Polyploid cells have genomes that contain multiples of the typical diploid chromosome number and are found in many different organisms. Studies in a variety of animal and plant developmental systems have revealed evolutionarily conserved mechanisms that control the generation of polyploidy and have recently begun to provide clues to its physiological function. These studies demonstrate that cellular polyploidy plays important roles during normal development and also contributes to human disease, particularly cancer