Abstract

Size control is essential for all proliferating cells, and is thought to be regulated by checkpoints that couple cell size to cell cycle progression. The aberrant cell-size phenotypes caused by mutations in the retinoblastoma (RB) tumor suppressor pathway are consistent with a role in size checkpoint control, but indirect effects on size caused by altered cell cycle kinetics are difficult to rule out. The multiple fission cell cycle of the unicellular alga Chlamydomonas reinhardtii uncouples growth from division, allowing direct assessment of the relationship between size phenotypes and checkpoint function. Mutations in the C. reinhardtii RB homolog encoded by MAT3 cause supernumerous cell divisions and small cells, suggesting a role for MAT3 in size control. We identified suppressors of an mat3 null allele that had recessive mutations in DP1 or dominant mutations in E2F1, loci encoding homologs of a heterodimeric transcription factor that is targeted by RB-related proteins. Significantly, we determined that the dp1 and e2f1 phenotypes were caused by defects in size checkpoint control and were not due to a lengthened cell cycle. Despite their cell division defects, mat3, dp1, and e2f1 mutants showed almost no changes in periodic transcription of genes induced during S phase and mitosis, many of which are conserved targets of the RB pathway. Conversely, we found that regulation of cell size was unaffected when S phase and mitotic transcription were inhibited. Our data provide direct evidence that the RB pathway mediates cell size checkpoint control and suggest that such control is not directly coupled to the magnitude of periodic cell cycle transcription.

Highlights

  • Size control is a fundamental property of proliferating cells, but the mechanisms that maintain cell size are still poorly defined [1]

  • We found that dp1 null mutants had a large-cell phenotype caused by an increased Commitment threshold size and a failure to initiate enough rounds of division during S phases and mitoses (S/M), opposite to the size checkpoint phenotypes of mat3 mutants

  • Isolation and Characterization of mat3 Bypass Suppressors To investigate the downstream targets of MAT3 that are required for regulation of cell division, we carried out an insertional mutagenesis screen for bypass suppressors of a null allele, mat3–4 [25,32]

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Summary

Introduction

Size control is a fundamental property of proliferating cells, but the mechanisms that maintain cell size are still poorly defined [1]. There is evidence for cell size checkpoint control [3,4,5,6], but the nature and existence of size checkpoints in animals are still debated [7]. There appears to be a fundamental relationship between cell size and DNA content that is likely to be involved in governing cell size, but the nature of that relationship is still unknown [8]. Cell size appears subject to input from multiple genetic pathways whose relative contributions to a size checkpoint mechanism are still unclear

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