Biology subjects

Brachova, P.

Publications and source records attributed to Brachova, P..

4 recordsLinked to original sources

bioRxiv2025

Dysregulation of alternative splicing patterns in the ovaries of reproductively aged mice

Female reproductive aging is characterized by progressive deterioration of ovarian function, yet the molecular mechanisms driving these changes remain incompletely understood. Here, we used long-read direct RNA-sequencing to map transcript isoform changes in mouse ovaries across reproductive age. Comparing young and aged mice after controlled gonadotropin stimulation, we identified widespread alternative splicing changes, including shifts in exon usage, splice site selection, and transcript boundaries. Aged ovaries exhibited increased isoform diversity, favoring distal start and end sites, and a significant rise in exon skipping and intron retention events. Many of these age-biased splicing events altered open reading frames, introduced premature stop codons, or disrupted conserved protein domains. Notably, mitochondrial genes were disproportionately affected. We highlight Ndufs4, a mitochondrial Complex I subunit, as a case in which aging promotes the alternative splicing of a truncated isoform lacking the canonical Pfam domain. Structural modeling suggests this splice variant could impair Complex I function, resulting in increased ROS production. Our data suggest a mechanistic link between splicing and mitochondrial dysfunction in the aging ovary. These findings support the model of the splicing-energy-aging axis in ovarian physiology, wherein declining mitochondrial function and adaptive or maladaptive splicing changes are intertwined. Our study reveals that alternative splicing is not merely a byproduct of aging but a dynamic, transcriptome-wide regulatory layer that may influence ovarian longevity. These insights open new avenues for investigating post-transcriptional mechanisms in reproductive aging and underscore the need to consider isoform-level regulation in models of ovarian decline. In briefReproductive aging is associated with changes in alternative splicing patterns in the mouse ovary. Our study identifies alterations in exon usage that may have altered protein function and important roles in ovarian physiology as well as in reproductive aging.

cell biology↗

bioRxiv2021

Codon composition in human oocytes reveals age-associated defects in mRNA decay

Oocytes from women of advanced reproductive age have lower developmental potential, yet the underlying mechanisms of this phenomena are incompletely understood. Oocyte maturation is dependent upon translational control of stored maternal mRNA that were synthesized during oocyte growth. We observed that GC content of mRNA was negatively associated with half-life in oocytes from reproductively young women (< 30 years), contrastingly directly with oocytes from reproductively aged women ([≥] 40 years) where mRNA half-lives were positively associated with GC nucleotide content. Additionally, we observed that mRNA half-lives were negatively associated with protein abundance in young oocytes, while GC content was positively associated with protein abundance in aged oocytes. Examination of codon composition during the GV-to-MII transition revealed that codons that facilitate rapid translation promoted mRNA stability and are considered optimal, while codons that slow translation destabilized mRNA, and are considered non-optimal. GC-containing codons were more optimal in reproductive aging, and also correlated positively with protein abundance. This study indicates that reproductive aging coincides with the stabilization of a subset of mRNA that have the potential to be over-translated during oocyte maturation, this is likely to lead to observed decreases in oocyte quality in older women. Because oocyte mRNA decay is translationally linked, this suggests that maternal aging causes defects in translation, which results in reduced translational efficiency and the retention of maternal mRNA that are normally degraded in oocytes from young women. In the case of oocytes, defects in translation can alter the RNA decay pathways and result in incorrect maternal mRNA dosage, which may negatively impact embryonic development.

developmental biology↗

bioRxiv2020

Loss of Cnot6l impairs inosine RNA modifications in mouse oocytes

Mammalian oocytes must degrade maternal transcripts through a process called translational mRNA decay, in which maternal mRNA undergoes translational activation, followed by deadenylation and mRNA decay. Once a transcript is translationally activated, it becomes deadenylated by the CCR4-NOT complex. Knockout of Cnot6l, a deadenylase within the CCR4-NOT complex, results in mRNA decay defects during MI entry. Knockout of Btg4, an adaptor protein of the CCR4-NOT complex, results in mRNA decay defects following fertilization. Therefore, mechanisms controlling mRNA turnover have significant impacts on oocyte competence and early embryonic development. Post-transcriptional inosine RNA modifications can impact mRNA stability, possibly through a translation mechanism. Here, we assessed inosine RNA modifications in oocytes from Cnot6l-/- and Btg4-/- mice, which display stabilization of mRNA and over-translation of the stabilized transcripts. If inosine modifications have a role in modulating RNA stability, we hypothesize that in these mutant backgrounds, we would observe changes or a disruption in inosine mRNA modifications. To test this, we used a computational approach to identify inosine RNA modifications in total and polysomal RNA-seq data during meiotic maturation (GV, MI, and MII stages). We observed pronounced depletion of inosine mRNA modifications in oocytes from Cnot6l-/-, but not in Btg4-/- mice. Additionally, analysis of ribosome-associated RNA revealed clearance of inosine modified mRNA. These observations suggest a novel mechanism of mRNA clearance during oocyte maturation, in which inosine-containing transcripts decay in an independent, but parallel mechanism to CCR4-NOT deadenylation.

developmental biology↗

bioRxiv2018

ADAR1 A-to-I RNA alters codon usage

BackgroundFully grown mammalian oocytes and eggs are transcriptionally quiescent, and therefore have a unique RNA environment in which cellular processes depend on post-transcriptional regulation. RNA editing of adenosines into inosines (A-to-I) by adenosine deaminases acting on RNA (ADARs) is a common post-transcriptional gene regulatory mechanism, yet it has not been systematically studied in oocytes.\n\nResultsA genome-wide RNA editing analysis of transcriptionally active growing oocytes from postnatal day 12 (PND12) mice, fully grown germinal vesicle (GV) oocytes, and transcriptionally quiescent metaphase II (MII) eggs indicates an abundant amount of A-to-I editing of mRNA transcripts. Editing of mRNA was greatest in GV oocyte and MII eggs compared to the PND12 immature oocytes, this was consistent with ADAR1 levels within these cells. Compared to somatic tissues, oocytes exhibited a different pattern of RNA editing, with a high proportion of RNA edits occurring in the coding regions. These edits resulted in nucleotide substitutions that were enriched at the third nucleotide of the codon (wobble position). Codon usage can affect mRNA stability and translation efficiency.\n\nConclusionsRNA editing in mouse oocytes is distinct from RNA editing in somatic cells due to increased frequencies of coding sequence RNA edits. We provide evidence in support of a previously unreported phenomenon of selective ADAR1 editing of the codon wobble position. Editing of the wobble position has the potential to fine tune post-transcriptional gene regulation through altering codon usage. This important observation advances our current understanding of RNA editing in mammalian cells.

developmental biology↗