Co-translational protein targeting facilitates centrosomal recruitment of PCNT during centrosome maturation in vertebrates

Abstract

As microtubule-organizing centers of animal cells, centrosomes guide the formation of the bipolar spindle that segregates chromosomes during mitosis. At mitosis onset, centrosomes maximize microtubule-organizing activity by rapidly expanding the pericentriolar material (PCM). This process is in part driven by the large PCM protein pericentrin (PCNT), as its level increases at the PCM and helps recruit additional PCM components. However, the mechanism underlying the timely centrosomal enrichment of PCNT remains unclear. Here, we show that PCNT is delivered co-translationally to centrosomes during early mitosis by cytoplasmic dynein, as evidenced by centrosomal enrichment of PCNT mRNA, its translation near centrosomes, and requirement of intact polysomes for PCNT mRNA localization. Additionally, the microtubule minus-end regulator, ASPM, is also targeted co-translationally to mitotic spindle poles. Together, these findings suggest that co-translational targeting of cytoplasmic proteins to specific subcellular destinations may be a generalized protein targeting mechanism.

Inter-dependent Centrosomal Co-localization of the cen and ik2 cis-Natural Antisense mRNAs in Drosophila

“…By contrast, exposure to harringtonine led to a marked disruption of cen mRNA localization, which became diffusely distributed in the cytoplasm ( Figure 6C). We conclude that the cen mRNA can undergo localized translation at the level of centrosomes and that its localization is dependent on the maintenance of intact polysomes, similar to recent observations for centrosomally localized pericentrin (PCNT) mRNA in HeLa cells (Sepulveda et al, 2018).…”

“…Next, we sought to test whether the centrosomal localization of cen mRNA is dependent on intact translating polysomes by treating embryos with translation inhibitors. To address this question, we performed FISH analyses on embryos treated with emetine, which stalls translation during elongation and stabilizes polysomes, or with harringtonine, a polysome disruptor that blocks the initiation step of translation (Fresno et al, 1977;Sepulveda et al, 2018). Although emetine induced morphological aberrations in a small subset of embryos, the centrosomal localization pattern of cen mRNA was similar to untreated or vehicle (DMSO)-treated embryos, both during interphase and in mitosis ( Figure 6C).…”

Proximity labeling at non-centrosomal microtubule-organizing centers reveals VAB-10B and WDR-62 as distinct microtubule regulators

“…Given the central role we found for actin in anchoring non-centrosomal microtubules in the embryonic intestine, it will be necessary to further investigate the role of proximity interactors that also interact with actin. RNA binding proteins have not previously been shown to localize to ncMTOCs, however the existence and function of RNA at the centrosome has a long history and so this class of proteins is an interesting category for further investigation (Marshall & Rosenbaum 2000;Alliegro et al, 2006;Sepulveda et al, 2018).…”

The Emerging Role of ncRNAs and RNA-Binding Proteins in Mitotic Apparatus Formation

“…Gle1 is localized in the PCM, and its depletion significantly attenuates signal intensity of PCNT, a centrosomal scaffold protein, at the centrosome [44]. The mRNA of PCNT has been reported to accumulate in centrosomes awaiting translation [45], but whether Gle1 regulates PCNT mRNA to be translated locally at the centrosome remains to be clarified.…”