Monosome Formation during Translation Initiation Requires the Serine/Arginine-Rich Protein Npl3

Abstract

The yeast shuttling serine/arginine-rich protein Npl3 is required for the export of mRNAs and pre-60S ribosomal subunits from the nucleus to the cytoplasm. Here, we report a novel function of Npl3 in translation initiation. A mutation in its C terminus that prevents its dimerization (npl3⌬100) is lethal to cells and leads to translational defects, as shown by [35 S]methionine incorporation assays and a hypersensitivity to the translational inhibitor cycloheximide. Moreover, this Npl3 mutant shows halfmers in polysomal profiles that are indicative of defects in monosome formation. Strikingly, the loss of the ability of Npl3 to dimerize does not affect mRNA and pre-60S export. In fact, the mRNA and rRNA binding capacities of npl3⌬100 and wild-type Npl3 are similar. Intriguingly, overexpression of the dimerization domain of Npl3 disturbs dimer formation and results in a dominantnegative effect, reflected in growth defects and a halfmer formation phenotype. In addition, we found specific genetic interactions with the ribosomal subunit joining factors Rpl10 and eukaryotic translation initiation factor 5B/Fun12 and detected a substantially decreased binding of npl3⌬100 to the Rpl10-containing complex. These findings indicate an essential novel function for Npl3 in the cytoplasm, which supports monosome formation for translation initiation.

Knockout of the Hmt1p Arginine Methyltransferase in Saccharomyces cerevisiae Leads to the Dysregulation of Phosphate-associated Genes and Processes

“…There is a dimerization precedent in hnRNP protein Npl3p, where nuclear Hmt1p-mediated arginine methylation increases selfdimerization by ϳ2-fold (52,104). The dimerization domain of Npl3p is in the C-terminal SR-domain, from positions 276 -364, and loss of Npl3p dimerization results in defects in translation (105). We hypothesize that the function of the putative arginine methylation site on Pho4p could be like Npl3p, potentially increasing its homodimerization.…”

Npl3, a new link between RNA-binding proteins and the maintenance of genome integrity

“…5,6 Npl3 is a multifunctional protein that participates in a variety of RNArelated processes, including transcription, splicing, mRNA export, and translation. [7][8][9][10] Additionally, a recent report has provided evidence of an Npl3 role in preventing genome instability. 11 The function of Npl3 at transcribed genes is necessary to avoid R-loop formation between the nascent mRNA and the transcribed …”

Post‐translational modification directs nuclear and hyphal tip localization of Candida albicans mRNA‐binding protein Slr1

“…The arginine‐rich domain of ScNpl3 modulates its nucleocytoplasmic transport (Senger et al ., ; McBride et al ., ; Baierlein et al ., ). To test whether the R‐rich domain influences the nuclear localization of Slr1, we deleted parts of this domain in Slr1‐GFP.…”

“…D), in addition to the cytoplasmic localization of slr1‐6SA‐GFP, suggested that Slr1 might function in part in the cytoplasm. S. cerevisiae SR‐like protein Npl3 is required for monosome formation during translation initiation and co‐fractionates with polysomes (Windgassen et al ., ; Baierlein et al ., ). We, therefore, compared the migration of wildtype Slr1‐GFP with that of ribosomal protein Rps3 through sucrose density gradients (Fig.…”

“…These R‐rich domains are often targets for arginine methylation (Henry and Silver, ; McBride et al ., ; Ariyachet et al ., ). The most thoroughly studied yeast SR‐like protein, S. cerevisiae Npl3, has been implicated in many aspects of mRNA metabolism from transcription and splicing to mRNA export and translation (Kadowaki et al ., ; Lee et al ., ; Bucheli and Buratowski, ; Dermody et al ., ; Kress et al ., ; Baierlein et al ., ). Both ScNpl3 and its S. pombe ortholog, SpSrp2, are essential, whereas SpSrp1 is not essential (Bossie et al ., ; Gross et al ., ; Lutzelberger et al ., ).…”