Biology subjects

Bromfield, E. G.

Publications and source records attributed to Bromfield, E. G..

3 recordsLinked to original sources

bioRxiv2025

The functional maturation of mouse spermatozoa is underpinned by global remodeling of the cellular phosphoproteome

The functional maturation of mammalian spermatozoa is driven by modification of their intrinsic proteome as the cells transit the male (epididymal sperm maturation) and female reproductive tracts (capacitation). Here, high-resolution mass spectrometry was used to interrogate the central role that phosphoproteomic changes play in the functional remodeling of mouse spermatozoa. This strategy identified 14,586 site-specific phosphorylation events, including the phosphorylation of 573 proteins and dephosphorylation of 426 during epididymal maturation and additional phosphorylation changes in 211 proteins linked to capacitation. We identified over 300 kinases that putatively govern these events, including three novel kinases (STK33, HIPK4, and PAK1) implicated in acrosomal exocytosis. The functional relevance of these data was confirmed via the use of knockout mouse models, which demonstrated several phosphoproteins as being essential for sperm motility and fertilization capacity. These findings illustrate that large-scale phosphorylation remodeling occurs during sperm maturation with implications extending to novel means of fertility regulation. HIGHLIGHTSO_LISperm phosphoproteome is dramatically remodeled during post-testicular maturation C_LIO_LIIdentification of >14,000 site-specific phosphorylation events providing comprehensive insight into sperm cell signaling events associated with functional maturation C_LIO_LIMajor changes in the sperm phosphoproteome coincide with epididymal maturation whereas capacitation results in more modest changes C_LIO_LIIdentification of 343 novel kinases potentially important for conferring functional maturity to spermatozoa and demonstrated role for STK33, HIPK4 and PAK1 kinases C_LIO_LIKnockout mouse models of 23 genes provided in vivo validation, with loss of these proteins leading to pronounced defects in sperm motility and fertilization capacity C_LIO_LIAll data is available via our interactive ShinySpermPhospho application - https://reproproteomics.shinyapps.io/ShinySpermPhospho/ C_LI

cell biology↗

bioRxiv2024

Sub-chronic elevation in ambient temperature drives alterations to the sperm epigenome and accelerates early embryonic development in mice

Forecasted increases in the prevalence and severity of extreme weather events accompanying changes in climatic behavior pose potential risk to the reproductive capacity of humans and animals of ecological and agricultural significance. While several studies have revealed that heat stress induced by challenges such as testicular insulation can elicit a marked negative effect on the male reproductive system, and particularly the production of spermatozoa, less is known about the immediate impact on male reproductive function following sub-chronic whole-body exposure to elevated ambient temperature. To address this knowledge gap, we exposed unrestrained male mice to heat stress conditions that emulate a heat wave (daily cycle of 8_h at 35{degrees}C followed by 16 h at 25{degrees}C) for a period of seven days. Neither the testes or epididymides of heat exposed male mice exhibited evidence of gross histological change, and similarly, spermatozoa of exposed males retained their functionality and ability to support embryonic development. However, the embryos generated from heat exposed spermatozoa experienced pronounced changes in gene expression linked to acceleration of early embryo development, aberrant blastocyst hatching and increased fetal weight. Such changes were causally associated with an altered sperm small non-coding RNA (sncRNA) profile, such that these developmental phenotypes were recapitulated by microinjection of wild-type embryos sired by control spermatozoa with RNAs extracted from heat exposed spermatozoa. Such data highlight that even a relatively modest excursion in ambient temperature can affect male reproductive function and identify the sperm sncRNA profile as a particular point of vulnerability to this imposed environmental stress. Significance StatementThe fidelity of sperm production underpins successful reproduction yet is highly vulnerable to various forms of environmental challenge, including heat stress. Despite this knowledge, we lack a complete understanding of the immediate impact on male reproduction of whole-body exposure to elevated ambient temperatures such as those encountered during a heatwave. By experimentally emulating heatwave conditions, we demonstrate that the spermatozoa of exposed male mice accumulate changes in their small RNA profile that are causally linked to pronounced changes in embryonic gene expression, accelerated pre-implantation development, aberrant blastocyst hatching, and increased fetal weight. Such data highlight that even a relatively modest alteration in ambient temperature can affect male reproductive function, demonstrating the acute sensitivity of sperm small RNAs to environmental stress.

cell biology↗

bioRxiv2020

The multi-scale architecture of mammalian sperm flagella and implications for ciliary motility

Motile cilia are molecular machines used by a myriad of eukaryotic cells to swim through fluid environments. However, available molecular structures represent only a handful of cell types, limiting our understanding of how cilia are modified to support motility in diverse media. Here, we use cryo-focused ion beam milling-enabled cryo-electron tomography to image sperm flagella from three mammalian species. We resolve in-cell structures of centrioles, axonemal doublets, central pair apparatus, and endpiece singlets, revealing novel protofilament-bridging microtubule inner proteins throughout the flagellum. We present native structures of the flagellar base, which is crucial for shaping the flagellar beat. We show that outer dense fibers are directly coupled to microtubule doublets in the principal piece but not in the midpiece. Thus, mammalian sperm flagella are ornamented across scales, from protofilament-bracing structures rein-forcing microtubules at the nano-scale to accessory structures that impose micron-scale asymmetries on the entire assembly. Our structures provide vital foundations for linking molecular structure to ciliary motility and evolution.

cell biology↗