Roles for the RNA polymerase III regulator MAFR-1 in regulating sperm quality in Caenorhabditis elegans

Abstract

The negative regulator of RNA polymerase (pol) III mafr-1 has been shown to affect RNA pol III transcript abundance, lipid biosynthesis and storage, progeny output, and lifespan. We deleted mafr-1 from the Caenorhabditis elegans genome and found that animals lacking mafr-1 replicated many phenotypes from previous RNAi-based studies and discovered a new sperm-specific role. Utilizing a yeast two-hybrid assay, we discovered several novel interactors of MAFR-1 that are expressed in a sperm- and germline-enriched manner. In support of a role for MAFR-1 in the male germline, we found mafr-1 null males have smaller spermatids that are less capable in competition for fertilization; a phenotype that was dependent on RNA pol III activity. Restoration of MAFR-1 expression specifically in the germline rescued the spermatid-related phenotypes, suggesting a cell autonomous role for MAFR-1 in nematode male fertility. Based on the high degree of conservation of Maf1 activity across species, our study may inform similar roles for Maf1 and RNA pol III in mammalian male fertility.

Cellular responses to long-term phosphate starvation of fission yeast: Maf1 determines fate choice between quiescence and death associated with aberrant tRNA biogenesis

“…Prior studies of the lifespan-determining role of Maf1 highlight distinct effects in metazoa and yeast ( 49 ). Whereas a homozygous Maf1 knockout in female mice resulted in an increase in average lifespan ( 50 ), deletion of Maf1 did not affect the lifespan of C. elegans ( 51 ). The CLS of S. cerevisiae grown to stationary phase in 0.2% glucose was acutely curtailed in maf1 Δ cells (∼5% survival after 12 days) compared to wild-type cells (60% survival) ( 52 ).…”

RNA Polymerase III, Ageing and Longevity

“…In contrast, the loss of the Maf1 orthologue MAFR-1 in C. elegans through RNAi knockdown extended lifespan under normal and CR feeding, and increased stress resistance (Cai and Wei, 2016). More recently, the Curran lab employed a CRISPRbased approach to generate a mafr-1 null mutant and reported no difference in the lifespan relative to WT worms (Hammerquist and Curran, 2020), despite showing some phenotypic overlap (e.g., increased expression of Pol III transcripts and elevated FIGURE 1 | Conserved relationships between TORC1, RNA Polymerase III and lifespan. The growth-promoting, anabolic functions mediated by Pol III activation represent at least one mechanism downstream of TORC1 that shortens adult health and survival (Filer et al, 2017).…”

“…In contrast, the loss of the Maf1 orthologue MAFR-1 in C. elegans through RNAi knockdown extended lifespan under normal and CR feeding, and increased stress resistance ( Cai and Wei, 2016 ). More recently, the Curran lab employed a CRISPR- based approach to generate a mafr-1 null mutant and reported no difference in the lifespan relative to WT worms ( Hammerquist and Curran, 2020 ), despite showing some phenotypic overlap (e.g., increased expression of Pol III transcripts and elevated intracellular lipids) with the mafr-1 RNAi knockdown worms studied by Cai and colleagues ( Cai and Wei, 2015 , 2016 ). In Drosophila , dMaf1 inhibition during development in either the whole organism or the fat body increased larval growth rates and body size, and enhanced protein synthesis rates through increased insulin signalling ( Rideout et al, 2012 ), while over-expression in the adult gut moderately promoted longevity ( Filer et al, 2017 ).…”

Analysis of Caenorhabditis elegans Sperm Number, Size, Activation, and Mitochondrial Content

“…Additionally, several mitochondrial mutants negatively impact C. elegans fertility, although the exact mechanism of action of these pathways is unknown ( Jonassen et al , 2002 ; Grad and Lemire, 2004 ). Recent work from our lab has demonstrated that mafr-1 , a regulator of RNA polymerase III ( Hammerquist and Curran, 2020 ), and alh-6 and prdh-1 , enzymes in the mitochondrial proline catabolism pathway (Yen et al , 2020; Yen and Curran, 2021 ), are required for proper sperm function in C. elegans , and these roles may be conserved in mammalian models ( Bonhoure et al , 2015 ; Yen and Curran, 2021 ).…”

“…We recently developed and published methods to assess various metrics of spermatid quality in C. elegans ( Hammerquist and Curran, 2020 ; Yen and Curran, 2020; Yen et al , 2020). Some of the methods we present, albeit with added details and refined techniques, are largely unchanged from their original publication decades ago ( Ward et al , 1983 ; Shakes and Ward, 1989 ; LaMunyon and Ward, 1998 ), whereas others, specifically the quantification of spermatid mitochondrial fusion, were first developed in our lab ( Yen et al , 2020 ).…”