The Transcriptomes of Two Heritable Cell Types Illuminate the Circuit Governing Their Differentiation

Abstract

The differentiation of cells into distinct cell types, each of which is heritable for many generations, underlies many biological phenomena. White and opaque cells of the fungal pathogen Candida albicans are two such heritable cell types, each thought to be adapted to unique niches within their human host. To systematically investigate their differences, we performed strand-specific, massively-parallel sequencing of RNA from C. albicans white and opaque cells. With these data we first annotated the C. albicans transcriptome, finding hundreds of novel differentially-expressed transcripts. Using the new annotation, we compared differences in transcript abundance between the two cell types with the genomic regions bound by a master regulator of the white-opaque switch (Wor1). We found that the revised transcriptional landscape considerably alters our understanding of the circuit governing differentiation. In particular, we can now resolve the poor concordance between binding of a master regulator and the differential expression of adjacent genes, a discrepancy observed in several other studies of cell differentiation. More than one third of the Wor1-bound differentially-expressed transcripts were previously unannotated, which explains the formerly puzzling presence of Wor1 at these positions along the genome. Many of these newly identified Wor1-regulated genes are non-coding and transcribed antisense to coding transcripts. We also find that 5′ and 3′ UTRs of mRNAs in the circuit are unusually long and that 5′ UTRs often differ in length between cell-types, suggesting UTRs encode important regulatory information and that use of alternative promoters is widespread. Further analysis revealed that the revised Wor1 circuit bears several striking similarities to the Oct4 circuit that specifies the pluripotency of mammalian embryonic stem cells. Additional characteristics shared with the Oct4 circuit suggest a set of general hallmarks characteristic of heritable differentiation states in eukaryotes.

Fungal Morphogenesis

“…The difference in gene expression among cells within a community could be caused by stochastic factors, determinant factors (e.g., age, cell-cycle stage), or a response to the microenvironment created by gradients of signaling molecules and the extracellular matrix (Kerr et al 2002;Kim et al 2008;Shank et al 2011). For instance, stochastic expression of Wor1 determines white-opaque switch in Candida (Zordan et al 2006;Tuch et al 2010;Porman et al 2013), and adhesin Cfl1 in the extracellular matrix stimulates morphogenesis from yeast to hypha in Cryptococcus . The ability to have mixed yeast and hyphal morphotypes was recently shown to enhance Cryptococcus competitiveness in vitro (Phadke et al 2013).…”

Budding off: bringing functional genomics toCandida albicans

“…Most recent studies of gene expression have used RNA sequencing (RNA-Seq) to obtain greater sensitivity and to reduce bias in assayed gene targets. RNA-Seq studies have identified 600 novel coding and over 1000 noncoding transcripts in C. albicans, many of which are regulated by environment and cell state [97,98]. Transcriptional profiles have now been defined under a wide variety of conditions both in vitro and in vivo [99][100][101][102][103].…”

“…Transcriptional profiles have now been defined under a wide variety of conditions both in vitro and in vivo [99][100][101][102][103]. Profiling of unique phenotypic states, including white, opaque, gray and GUT (gastrointestinally induced transition), has been used to define the transcriptional networks regulating these states, and has revealed that these networks resemble those controlling cell fate in metazoans [34,75,98,104].…”

“…For example, experiments have revealed the widespread presence of antisense transcripts and untranslated regions of varying lengths, dependent both on cell state and environmental conditions [97,98,105]. In the case of the white-opaque switch, RNA-Seq analysis of sense-antisense transcript pairs showed a significant anti-correlation, suggesting that many antisense transcripts act to repress their sense counterparts [98].…”

“…For example, experiments have revealed the widespread presence of antisense transcripts and untranslated regions of varying lengths, dependent both on cell state and environmental conditions [97,98,105]. In the case of the white-opaque switch, RNA-Seq analysis of sense-antisense transcript pairs showed a significant anti-correlation, suggesting that many antisense transcripts act to repress their sense counterparts [98]. Antisense transcripts also regulate the yeastto-hyphal switch; the Brg1 transcription factor induces an antisense transcript against the NRG1 sense transcript, thereby targeting the sense transcript for destruction and stimulating hypha-specific gene expression [106].…”