RESUMO
Transient exposures to environmental stresses induce altered physiological states in exposed cells that persist after the stresses have been removed. These states, referred to as cellular memory, can even be passed on to daughter cells and may thus be thought of as embodying a form of epigenetic inheritance. We find that meiotically produced spores in the budding yeast S. cerevisiae possess a state of heightened stress resistance that, following their germination, persists for numerous mitotic generations. As yeast meiotic development is essentially a starvation response that a/alpha diploid cells engage, we sought to model this phenomenon by subjecting haploid cells to starvation conditions. We find also that haploid cells exposed to glucose withdrawal acquire a state of elevated stress resistance that persists after the reintroduction of these cells to glucose-replete media. Following release from lengthy durations of glucose starvation, we confirm that this physiological state of enhanced stress resistance is propagated in descendants of the exposed cells through two mitotic divisions before fading from the population. In both haploid starved cells and diploid produced meiotic spores we show that their cellular memories are not attributable to trehalose, a widely regarded stress protectant that accumulates in these cell types. Moreover, the transiently heritable stress resistant state induced by glucose starvation in haploid cells is independent of the Msn2/4 transcription factors, which are known to program cellular memory induced by exposure of cells to NaCl. Our findings identify new developmentally and nutritionally induced states of cellular memory that exhibit striking degrees of persistence and mitotic heritability.
RESUMO
MicroRNAs (miRNAs) comprise a class of short noncoding RNA molecules that play diverse developmental and physiological roles by controlling mRNA abundance and protein output of the vast majority of transcripts. Despite the importance of miRNAs in regulating gene function, we still lack a complete understanding of how miRNAs themselves are transcriptionally regulated. To fill this gap, we predicted regulatory sequences by searching for abundant short motifs located upstream of miRNAs in eight species of Caenorhabditis nematodes. We identified three conserved motifs across the Caenorhabditis phylogeny that show clear signatures of purifying selection from comparative genomics, patterns of nucleotide changes in motifs of orthologous miRNAs, and correlation between motif incidence and miRNA expression. We then validated our predictions with transgenic green fluorescent protein reporters and site-directed mutagenesis for a subset of motifs located in an enhancer region upstream of let-7 We demonstrate that a CT-dinucleotide motif is sufficient for proper expression of GFP in the seam cells of adult C. elegans, and that two other motifs play incremental roles in combination with the CT-rich motif. Thus, functional tests of sequence motifs identified through analysis of molecular evolutionary signatures provide a powerful path for efficiently characterizing the transcriptional regulation of miRNA genes.
