SUS1 introns are required for efficient mRNA nuclear export in yeast

Abstract

Efficient coupling between mRNA synthesis and export is essential for gene expression. Sus1/ENY2, a component of the SAGA and TREX-2 complexes, is involved in both transcription and mRNA export. While most yeast genes lack introns, we previously reported that yeast SUS1 bears two. Here we show that this feature is evolutionarily conserved and critical for Sus1 function. We determine that while SUS1 splicing is inefficient, it responds to cellular conditions, and intronic mutations either promoting or blocking splicing lead to defects in mRNA export and cell growth. Consistent with this, we find that an intron-less SUS1 only partially rescues sus1Δ phenotypes. Remarkably, splicing of each SUS1 intron is also affected by the presence of the other and by SUS1 exonic sequences. Moreover, by following SUS1 RNA and protein levels we establish that nonsense-mediated decay (NMD) pathway and the splicing factor Mud2 both play a role in SUS1 expression. Our data (and those of the accompanying work by Hossain et al.) provide evidence of the involvement of splicing, translation, and decay in the regulation of early events in mRNP biogenesis; and imply the additional requirement for a balance in splicing isoforms from a single gene.

An intronic RNA structure modulates expression of the mRNA biogenesis factor Sus1

“…5) suggests that the reduced amount of Sus1 protein likely produced by these mutants is still enough to partially complement Sus1 functions, including sus1Δ temperature sensitive growth defect (data not shown). This is interesting, since overexpression of Sus1 has been shown to largely impair its functionality (Cuenca-Bono et al 2011;Hossain et al 2011). …”

“…Fluorescent in situ hybridization (FISH) against poly(A) + RNA was done as described by Cuenca-Bono et al (2011); yeast cells were grown in 100 mL of SC-Leu medium at 30°C to an 0.3 OD600. Then, cultures were rapidly shifted to 37°C incubator for 2 h. After hybridization, slides were mounted using VECTASHIELD Mounting Medium with DAPI.…”

“…Previous analyses with wild-type cells incubated at 42°C revealed reduction of SUS1 expression, accompanied by accumulation of unspliced SUS1 transcripts and decrease of fully spliced mRNA (Cuenca-Bono et al 2011). We tested the effect of the different structural mutants in these conditions by semi-q-RT-PCR (Fig.…”

“…The functional relevance of the presence of two introns in the SUS1 pre-mRNA has been studied previously (Cuenca-Bono et al 2011;Hossain et al 2011). While intron 2 (I2) has consensus splice sequences, intron 1 (I1) is characterized by the presence of nonconsensus 5 ′ splice site (SS) (GUAUGA instead of canonical GUAUGU) and branch site (BS) (UACUGAC instead of UACUAAC).…”

IDENTIFICATION OF RNA STRUCTURES MODULATING THE EXPRESSION OF THE mRNA BIOGENESIS FACTOR SUS1

“…cerevisiae genes are intronless, the SUS1 gene consists of three exons (of 71, 140 and 77 nt) and two introns (of 80 and 70 nt), and its splicing and expression levels are regulated by different mechanisms [1], [52]. [53], [54].…”