Kinetochore inactivation by expression of a repressive mRNA

Abstract

Differentiation programs such as meiosis depend on extensive gene regulation to mediate cellular morphogenesis. Meiosis requires transient removal of the outer kinetochore, the complex that connects microtubules to chromosomes. How the meiotic gene expression program temporally restricts kinetochore function is unknown. We discovered that in budding yeast, kinetochore inactivation occurs by reducing the abundance of a limiting subunit, Ndc80. Furthermore, we uncovered an integrated mechanism that acts at the transcriptional and translational level to repress NDC80 expression. Central to this mechanism is the developmentally controlled transcription of an alternate NDC80 mRNA isoform, which itself cannot produce protein due to regulatory upstream ORFs in its extended 5’ leader. Instead, transcription of this isoform represses the canonical NDC80 mRNA expression in cis, thereby inhibiting Ndc80 protein synthesis. This model of gene regulation raises the intriguing notion that transcription of an mRNA, despite carrying a canonical coding sequence, can directly cause gene repression.

NF-Y controls fidelity of transcription initiation at gene promoters through maintenance of the nucleosome-depleted region

“…This was surprising considering that changes in transcript levels generally result in proportional changes in protein abundance (Spearman correlation of ~0.8 34,35 ). Recent studies have shown that upstream ORFs (uORFs) can have either a favourable or a deleterious effect on downstream mRNA translation [36][37][38][39][40] and suggested that the abundance of transcript is less important than the difference in translatability of the canonical versus ectopic transcript 38 . Thus, in situations wherein an upstream extension of the transcript leads to altered protein production, we speculate that this could be due to introduction of an uORF.…”

“…4d). Given that uORFs can modulate downstream translation and thus act as potent regulators of translation and protein expression [37][38][39][40]57 , it is conceivable that translation initiation from non-canonical start codon(s) within uORFs alters the reading frame and/or protein length; alternatively, it may affect the efficiency with which ribosomes translate the rest of the transcript 58 .…”

Tunable Transcriptional Interference at the Endogenous Alcohol Dehydrogenase Gene Locus inDrosophila melanogaster

“…In addition to non-coding RNAs, mRNA isoforms have also been linked to transcriptional interference. For genes with more than one promoter, transcription from the distal promoter may not only produce a distinct mRNA isoform, but could also lead to the repression of an mRNA isoform transcribed from the open reading frame (ORF)-proximal gene promoter (Corbin and Maniatis, 1989;Moseley et al, 2002;Sehgal et al, 2008;Liu et al, 2015;Chen et al, 2017). In addition, since distinct mRNA isoforms may differ in their translational efficiency, regulation of promoter choice may impact gene expression at the protein level.…”

“…In addition, since distinct mRNA isoforms may differ in their translational efficiency, regulation of promoter choice may impact gene expression at the protein level. In some instances, this difference in translational efficiency is due to the presence of upstream ORFs (uORFs) in the 5' leader of the distal promoter-derived mRNA isoform, which could inhibit translation of the protein-coding ORF (Moseley et al, 2002;Law et al, 2005;Sehgal et al, 2008;Ingolia et al, 2011;Rojas-Duran and Gilbert, 2012;Brar et al, 2012;Chew et al, 2016;Bird and Labbé, 2017;Chen et al, 2017;Cheng et al, 2018;Zhang et al, 2018) . As a result, in these cases, transcription of a distal promoter-derived mRNA isoform causes downregulation of protein expression through the integration of two seemingly disparate mechanisms of transcriptional and translational repression Cheng et al, 2018;Van Dalfsen et al, 2018;Hollerer et al submitted).…”

Aurora B-dependent Ndc80 Degradation Regulates Kinetochore Composition in Meiosis

“…When the ∆2-28 or ∆11-19 mutant were combined with a 425 mutant that prematurely forms spindle microtubules in meiotic prophase (CUP-CLB3), 426we observed that sister chromatids, instead of homologous chromosomes, segregated 427in meiosis I (Figure 7E). The same phenotype occurs when Ndc80 is overexpressed in 428 meiotic prophase or when the LUTI-based repression of Ndc80 synthesis is disrupted 429(Miller et al 2012;Chen et al 2017). Therefore, the lack of Ndc80 turnover in meiotic 430…”