Functional Dissection of the Nascent Polypeptide-Associated Complex in Saccharomyces cerevisiae.

ABSTRACT: Both the yeast nascent polypeptide-associated complex (NAC) and the Hsp40/70-based chaperone system RAC-Ssb are systems tethered to the ribosome to assist cotranslational processes such as folding of nascent polypeptides. While loss of NAC does not cause phenotypic changes in yeast, the simultaneous deletion of genes coding for NAC and the chaperone Ssb (nac?ssb?) leads to strongly aggravated defects compared to cells lacking only Ssb, including impaired growth on plates containing L-canavanine or hygromycin B, aggregation of newly synthesized proteins and a reduced translational activity due to ribosome biogenesis defects. In this study, we dissected the functional properties of the individual NAC-subunits (?-NAC, ?-NAC and ?'-NAC) and of different NAC heterodimers found in yeast (??-NAC and ??'-NAC) by analyzing their capability to complement the pleiotropic phenotype of nac?ssb? cells. We show that the abundant heterodimer ??-NAC but not its paralogue ??'-NAC is able to suppress all phenotypic defects of nac?ssb? cells including global protein aggregation as well as translation and growth deficiencies. This suggests that ??-NAC and ??'-NAC are functionally distinct from each other. The function of ??-NAC strictly depends on its ribosome association and on its high level of expression. Expression of individual ?-NAC, ?'-NAC or ?-NAC subunits as well as ??'-NAC ameliorated protein aggregation in nac?ssb? cells to different extents while only ?-NAC was able to restore growth defects suggesting chaperoning activities for ?-NAC sufficient to decrease the sensitivity of nac?ssb? cells against L-canavanine or hygromycin B. Interestingly, deletion of the ubiquitin-associated (UBA)-domain of the ?-NAC subunit strongly enhanced the aggregation preventing activity of ??-NAC pointing to a negative regulatory role of this domain for the NAC chaperone activity in vivo.

SUBMITTER: Ott AK 

PROVIDER: S-EPMC4664479 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature