Graphical Abstract Highlights d Tumorigenesis depends on functional OXPHOS d OXPHOS-derived ATP is not required for tumor formation d DHODH-driven pyrimidine biosynthesis requires CoQ redoxcycling d CoQ redox-cycling via OXPHOS drives tumorigenesis through pyrimidine biosynthesis

Recently, we showed that generation of tumours in syngeneic mice by cells devoid of mitochondrial (mt) DNA (ρ0 cells) is linked to the acquisition of the host mtDNA. However, the mechanism of mtDNA movement between cells remains unresolved. To determine whether the transfer of mtDNA involves whole mitochondria, we injected B16ρ0 mouse melanoma cells into syngeneic C57BL/6Nsu9-DsRed2 mice that express red fluorescent protein in their mitochondria. We document that mtDNA is acquired by transfer of whole mitochondria from the host animal, leading to normalisation of mitochondrial respiration. Additionally, knockdown of key mitochondrial complex I (NDUFV1) and complex II (SDHC) subunits by shRNA in B16ρ0 cells abolished or significantly retarded their ability to form tumours. Collectively, these results show that intact mitochondria with their mtDNA payload are transferred in the developing tumour, and provide functional evidence for an essential role of oxidative phosphorylation in cancer.DOI: http://dx.doi.org/10.7554/eLife.22187.001

Chemical communication is mediated by signal production and signal perception and in house mice (Mus musculus), both processes involve lipocalin proteins (OBP, MUP, LCN) that transport volatiles and protect them in tissues where they are produced. However, potential roles of lacrimal, nasal, and salivary lipocalins are still not well known. We aimed to determine the expression of the recently described family of odorant binding proteins (Obp), along with major urinary proteins (Mup) across different tissues in wild mice (Mus musculus) to assess the importance of these proteins based on their quantity in particular expression sites. We performed qPCR analysis of selected Mup, Lcn, Obp genes, and predicted Obp members to study their expression in selected tissues. We identified new members of the mouse odorant binding protein gene family in two subspecies, M. m. musculus and M. m. domesticus. We show that Mup4 and Mup5 from the phylogenetically older group-A are co-expressed with Obps in orofacial tissues. We also identified a sexually dimorphic pattern of female-biased Obp7 and male-biased Mup4 expression in lacrimal glands. OBPs, MUPs, and LCNs are produced in parallel, which may function to widen the spectrum of bound ligands, potentially including the degradation products of olfactory signals and/or toxic compounds. Moreover, our study demonstrates that several pheromone transporters from the lipocalin family are co-expressed in the nasal and lacrimal tissues of mice with the newly detected OBPs that further expand the already diverse mouse lipocalin family.

Sperm chromatin reveals two characteristic features in that protamines are the predominant nuclear proteins and remaining histones are highly acetylated. histone h4 acetylated at lysine 12 (h4K12ac) is localized in the post-acrosomal region, while protamine-1 is present within the whole nucleus. chromatin immunoprecipitation in combination with promoter array analysis allowed genome-wide identification of h4K12ac binding sites. previously, we reported enrichment of h4K12ac at cTcF binding sites and promoters of genes involved in developmental processes. here, we demonstrate that h4K12ac is enriched predominantly between ± 2 kb from the transcription start site. In addition, we identified developmentally relevant h4K12ac-associated promoters with high expression levels of their transcripts stored in mature sperm. The highest expressed mRNa codes for testis-specific phD finger protein-7 (phF7), suggesting an activating role of h4K12ac in the regulatory elements of this gene. h4K12ac-associated genes revealed a weak correlation with genes expressed at 4-cell stage human embryos, while 23 h4K12ac-associated genes were activated in 8-cell embryo and 39 in the blastocyst. Genes activated in 4-cell embryos are involved in gene expression, histone fold and DNadependent transcription, while genes expressed in the blastocyst were classified as involved in developmental processes. Immunofluorescence staining detected h4K12ac from the murine male pronucleus to early stages of embryogenesis. aberrant histone acetylation within developmentally important gene promoters in infertile men may reflect insufficient sperm chromatin compaction, which may result in inappropriate transfer of epigenetic information to the oocyte.

There is pronounced promiscuity and sperm competition in long-tailed field mice (Apodemus sylvaticus). These mice have evolved unusual sperm behaviour favouring rapid fertilisation, including dynamic formation of sperm trains and their subsequent dissociation. The cell surface complement regulatory (CReg) protein CD46 is broadly expressed in eutherian mammals other than rodents, in which it is expressed solely on the spermatozoal acrosomal membrane. Ablation of the CD46 gene has been associated with a faster acrosome reaction (AR) rate in inbred laboratory mice. Here, we demonstrate that wild-caught field mice of three species, A. sylvaticus, A. flavicollis and A. microps, exhibit a more rapid AR than wild-caught house mice Mus musculus or inbred laboratory BALB/c mice. We also demonstrate that wild-caught field mice of these three species, unlike house mice, produce alternatively spliced transcripts of testicular CD46 mRNA lacking exons 5-7 or 6-7, together with an extended 3 0 -and often truncated 5 0 -utr, leading to failure to express any sperm CD46 protein in both the testis and epididymis. Male field mice may therefore have traded expression of this CReg protein for acrosomal instability, providing a novel genus-specific strategy to favour rapid fertilisation and competitive advantage in the promiscuous reproductive behaviour of wild field mice.

In vitro spermatogenesis (IVS) has already been successfully achieved in rodents by organotypic and soft matrix culture systems. However, the former does not allow single cell input, and the latter presents as a simple thick layer in which all cells are embedded. We explored a new culture system using a mouse model by employing an alginate-based hydrogel and 3D bioprinting, to control scaffold design and cell deposition. We produced testicular constructs consisting of printed cell-free scaffolds (CFS) with prepubertal testicular cells (TC) in their easy-to-access macropores. Here, the pores represented the only cell compartment (TC/CFS). Double-cell compartment testicular constructs were achieved by culturing magnetic-activated cell sorting-enriched epithelial cells in the pores of interstitial cell-laden scaffolds (CD49f + /CLS). Cell spheres formed in the pores in the weeks following cell seeding on both CFS and CLS. Although restoration of the tubular architecture was not observed, patches of post-meiotic cells including elongated spermatids were found in 66% of TC/CFS. Differentiation up to the level of round spermatids and elongated spermatids was observed in all and 33% of CD49f + /CLS constructs, respectively. Organ culture served as the reference method for IVS, with complete spermatogenesis identified in 80% of cultivated prepubertal tissue fragments. So far, this is the first report applying a 3D bioprinting approach for IVS. Further optimization of the scaffold design and seeding parameters might be permissive for tubular architecture recreation and thereby increase the efficiency of IVS in printed testicular constructs. While it remains to be tested whether the gametes generated on the alginate-based scaffolds can support embryogenesis following IVF, this IVS approach might be useful for (patho)physiological studies and drug-screening applications.

The mechanism of gamete fusion remains largely unknown on a molecular level despite its indisputable significance. Only a few of the molecules required for membrane interaction are known, among them IZUMO1, which is present on sperm, tetraspanin CD9, which is present on the egg, and the newly found oolema protein named Juno. A concept of a large multiprotein complex on both membranes forming fusion machinery has recently emerged. The Juno and IZUMO1, up to present, is the only known extracellular receptor pair in the process of fertilization, thus, facilitating the essential binding of gametes. However, neither IZUMO1 nor Juno appears to be the fusogenic protein. At the same time, the tetraspanin is expected to play a role in organizing the egg membrane order and to interact laterally with other factors. This review summarizes, to present, the known molecules involved in the process of sperm-egg fusion. The complexity and expected redundancy of the involved factors makes the process an intricate and still poorly understood mechanism, which is difficult to comprehend in its full distinction.

The crucial role that oestrogens play in male reproduction has been generally accepted; however, the exact mechanism of their action is not entirely clear and there is still much more to be clarified. The oestrogen response is mediated through oestrogen receptors, as well as classical oestrogen receptors’ variants, and their specific co-expression plays a critical role. The importance of oestrogen signalling in male fertility is indicated by the adverse effects of selected oestrogen-like compounds, and their interaction with oestrogen receptors was proven to cause pathologies. The aims of this review are to summarise the current knowledge on oestrogen signalling during spermatogenesis and sperm maturation and discuss the available information on oestrogen receptors and their splice variants. An overview is given of species-specific differences including in humans, along with a detailed summary of the methodology outcome, including all the genetically manipulated models available to date. This review provides coherent information on the recently discovered mechanisms of oestrogens’ and oestrogen receptors’ effects and action in both testicular somatic and germ cells, as well as in mature sperm, available for mammals, including humans.