Potential RNA Binding Proteins in Saccharomyces cerevisiae Identified as Suppressors of Temperature-Sensitive Mutations in NPL3

Abstract

The NPL3 gene of the yeast Saccharomyces cerevisiae encodes a protein with similarity to heterogeneous nuclear ribonucleoproteins (hnRNPs). Npl3p has been implicated in many nuclear-related events including RNA export, protein import, and rRNA processing. Several temperature-sensitive alleles of NPL3 have been isolated. We now report the sequence of these alleles. For one allele, npl3-1, four complementation groups of suppressors have been isolated. The cognate genes for the two recessive mutants were cloned. One of these is the previously known RNA15, which, like NPL3, also encodes a protein with similarity to the vertebrate hnRNP A/B protein family. The other suppressor corresponds to a newly defined gene we term HRP1, which also encodes a protein with similarity to the hnRNP A/B proteins of vertebrates. Mutations in HRP1 suppress all npl3 temperature-sensitive alleles but do not bypass an npl3 null allele. We show that HRP1 is essential for cell growth and that the corresponding protein is located in the nucleus. The discovery of two hnRNP homologues that can partially suppress the function of Npl3p, also an RNA binding protein, will be discussed in terms of the possible roles for Npl3p in RNA metabolism.

A Distinct and Parallel Pathway for the Nuclear Import of an mRNA-binding Protein

“…Deletion of NPL3 leads to inviability or very severe growth defects, depending on the yeast strain investigated. Npl3p shuttles in and out of the nu- cleus and has structural similarities to other RNA-binding proteins (Russell and Tollervey, 1992;Flach et al, 1994;Henry et al, 1996;Lee et al, 1996). These similarities include two central RNA recognition motifs flanked by a carboxy-terminal glycine-rich domain that contains a series of RGGY/F repeats (Wilson et al, 1994).…”

RNA Polymerase II C-Terminal Domain: Tethering Transcription to Transcript and Template

“…Npl3 copurifies with Pol II, interacts with the Pol II CTD, and is recruited to chromatin early in the transcription cycle and mutations of Npl3 that reduce RNA binding lead to a reduced elongation rate . After transcription termination Npl3 remains bound to the mRNA and facilitates its export to the cytoplasm . Once in the cytoplasm Npl3 plays a role in translation through interaction with ribosomal proteins and the mRNA poly­(A)-binding protein Pab1 .…”

Genome-Wide Distribution of Yeast RNA Polymerase II and Its Control by Sen1 Helicase

“…A 3-fold increase in the mRNA from HRP1 in the sen1 mutant strain confirmed Sen1-dependent attenuation (Figure 6C). S. cerevisiae Hrp1 (also called Nab4) is an mRNAbinding protein that shuttles between the nucleus and cytoplasm and influences poly(A) site selection (Henry et al, 1996;Kessler et al, 1997;Chen and Hyman, 1998;Minvielle-Sebastia et al, 1998). Hrp1 binds to AU dinucleotide repeat sequences (Kim Guisbert et al, 2005), which are present as efficiency elements in cleavage and polyadenylation signals (Guo and Sherman, 1996) and in Nrd1/Sen1-dependent snoRNA gene terminators (Steinmetz et al, 2006).…”