Regulated Antisense Transcription Controls Expression of Cell-Type-Specific Genes in Yeast

Abstract

Transcriptome profiling studies have recently uncovered a large number of noncoding RNA transcripts (ncRNAs) in eukaryotic organisms, and there is growing interest in their role in the cell. For example, in haploid Saccharomyces cerevisiae cells, the expression of an overlapping antisense ncRNA, referred to here as RME2 (Regulator of Meiosis 2), prevents IME4 expression. In diploid cells, the a1-␣2 complex represses the transcription of RME2, allowing IME4 to be induced during meiosis. In this study we show that antisense transcription across the IME4 promoter region does not block transcription factors from binding and is not required for repression. Mutational analyses found that sequences within the IME4 open reading frame (ORF) are required for the repression mediated by RME2 transcription. These results support a model where transcription of RME2 blocks the elongation of the full-length IME4 transcript but not its initiation. We have found that another antisense transcript, called RME3, represses ZIP2 in a cell-type-specific manner. These results suggest that regulated antisense transcription may be a widespread mechanism for the control of gene expression and may account for the roles of some of the previously uncharacterized ncRNAs in yeast.One of the main paradigms for the control of gene expression is that regulatory proteins bind to the promoter regions of genes to activate or repress transcription. However, it is now clear that noncoding RNAs (ncRNAs) also play important roles in gene regulation. For example, RNA interference (RNAi)-mediated regulation controls gene expression in Caenorhabditis elegans, Arabidopsis thaliana, humans, and many other organisms (16,24). However, a large number of ncRNAs do not appear to be involved in RNAimediated regulation. For example, more than 900 ncRNAs are expressed in the yeast Saccharomyces cerevisiae (11,15,33,40,45). Several of these ncRNAs act to regulate gene expression in yeast (2,5,18,26). However, S. cerevisiae lacks the enzymes Dicer and Argonaute, which are required for RNAi, and therefore, it must utilize different mechanisms for ncRNA-mediated regulation (12). In this paper we investigate how two antisense ncRNAs regulate the expression of genes required for meiosis in yeast.Under starvation conditions, diploid yeast undergoes meiosis and sporulation to form four haploid spores. This process involves the expression of more than 500 genes that are highly regulated in a coordinated manner (7,28). Entry into the meiotic pathway is controlled by the expression of IME1, the master initiator of meiosis (20, 29). There are two signals that regulate IME1 expression (Fig. 1A). One signal relates to the nutritional status of the cell, activating IME1 expression when the cell is starved of both nitrogen and a fermentable carbon source (14, 37). The second signal operates through cell-typespecific regulation, which allows the expression of meiotic genes only in a/␣ diploid cells. Cell-type-specific regulation is controlled by the a1-␣2 repressor complex, which reg...

Investigating Host Induced Meiosis in a Fungal Plant Pathogen

“…Their findings were consistent with a transcriptional interference mechanism, since transcription of Rme2 in haploid cells only interfered with Ime4 sense expression in cis (Hongay et al, 2006). Similarly, Zip2, a meiosis-specific protein involved in synaptonemal complex formation, shows cell-type specific sense and antisense expression (Gelfand et al, 2011). Haploids express an antisense transcript to Zip2, called Rme3 (regulator of meiosis 3) and MAT a/ diploids express Zip2 sense RNA.…”

“…The a1-2 protein heterodimer silences expression of Rme3 in MAT a/ diploids, enabling expression of Zip2. Gelfand et al (2011) expanded on previous research to show that Rme2 extension through the Ime4 promoter region was not required for Ime4 repression and Rme3 does not extend through the entire Zip2 ORF, indicating that both Rme2 and Rme3 do not interfere with TATA-binding proteins or polymerase binding in the promoter regions. Additionally, a 450 bp region within Ime4 was essential for Rme2-mediated repression.…”

“…This suggested that the 450 bp region may only be transcribed in a single direction at one time, or that extension of the transcript is terminated by specific protein complexes which bind this region, or chromatin remodelling occurs at this site. Overall, transcriptional interference may be prevalent in controlling yeast gene expression (Gelfand et al, 2011). Such mechanisms may be useful in fine-tuning condition-specific gene expression, especially when "leaky" expression of certain genes may be harmful to the cell.…”

“…Hongay et al (2006) observed cell-type specific sense and antisense transcription at the Ime4 locus. Haploids expressed an antisense transcript to Ime4, called Rme2 (regulator of meiosis 2; Gelfand et al, 2011), and MAT a/ diploids expressed Ime4 sense RNA. In MAT a/ diploids, the a1-2 protein heterodimer silences expression of Rme2, enabling the expression of Ime4, full expression of Ime1, and entry into meiosis.…”

Budding Yeast for Budding Geneticists: A Primer on the Saccharomyces cerevisiae Model System

“…Additional genomic elements have been discovered that are able to regulate transcription of nearby genes through the generation of noncoding RNAs (e.g., see Hongay et al 2006;Houseley et al 2008;Pinskaya et al 2009;Gelfand et al 2011;van Werven et al 2012;Castelnuovo et al 2013), and, based on the finding that noncoding RNA synthesis appears to be a widespread phenomenon in budding yeast (David et al 2006;Neil et al 2009;Xu et al 2009), many more such regulatory examples are likely to be identified in the future.…”