During spermiogenesis, haploid round spermatids undergo dramatic cell differentiation and morphogenesis to give rise to mature spermatozoa for fertilization, including nuclear elongation, chromatin remodeling, acrosome formation, and development of flagella. The molecular mechanisms underlining these fundamental processes remain poorly understood. Here, we report that MNS1, a coiled-coil protein of unknown function, is essential for spermiogenesis. We find that MNS1 is expressed in the germ cells in the testes and localizes to sperm flagella in a detergent-resistant manner, indicating that it is an integral component of flagella. MNS1–deficient males are sterile, as they exhibit a sharp reduction in sperm production and the remnant sperm are immotile with abnormal short tails. In MNS1–deficient sperm flagella, the characteristic arrangement of “9+2” microtubules and outer dense fibers are completely disrupted. In addition, MNS1–deficient mice display situs inversus and hydrocephalus. MNS1–deficient tracheal motile cilia lack some outer dynein arms in the axoneme. Moreover, MNS1 monomers interact with each other and are able to form polymers in cultured somatic cells. These results demonstrate that MNS1 is essential for spermiogenesis, the assembly of sperm flagella, and motile ciliary functions. Cilia are microtubule-based structures present in virtually all cells in vertebrates. Cilia have diverse functions in development, growth, signaling, and fertilization. Primary ciliary dyskinesia (PCD) affects one in 16,000 individuals. PCD is characterized by bronchiectasis and chronic sinusitis, and is often associated with situs inversus and male infertility. The genetic cause of PCD is heterogeneous. Some cases of PCD in humans and animals are caused by single genic mutations such as mutations in genes encoding microtubule-based dynein arm components. We have characterized a protein called MNS1 and found that it plays an essential role in ciliary functions in mice. MNS1 is a novel and integral component of sperm flagella. Mice lacking MNS1 exhibit male sterility as evidenced by abnormal assembly of sperm flagella. MNS1–deficient mice also display defects in left–right asymmetry patterning of internal organs and hydrocephalus. Therefore, mutations in MNS1 may contribute to male infertility and PCD in humans.

Polycomb group proteins mediate transcriptional silencing in diverse developmental processes. Sex chromosomes undergo chromosome-wide transcription silencing during male meiosis. Here we report that mouse SCML2 (Sex comb on midleg-like 2), an X chromosome-encoded polycomb protein, is specifically expressed in germ cells, including spermatogonia, spermatocytes, and round spermatids. SCML2 associates with phosphorylated H2AX and localizes to the XY body in spermatocytes. Loss of SCML2 in mice causes defective spermatogenesis, resulting in sharply reduced sperm production. SCML2 interacts with and recruits a deubiquitinase, USP7, to the XY body in spermatocytes. In the absence of SCML2, USP7 fails to accumulate on the XY body, whereas H2A monoubiquitination is dramatically augmented in the XY chromatin. Our results demonstrate that the SCML2/USP7 complex constitutes a novel molecular pathway in modulating the epigenetic state of sex chromosomes during male meiosis. Chromatin-modifying enzymes regulate the chromatin state during development and disease. Polycomb group proteins control the expression of homeotic genes in developmental patterning by catalyzing post-translational modifications of histones—core protein components of the chromatin. Most studies have focused on widely expressed polycomb proteins. However, the tissue-specific roles of polycomb proteins are poorly understood. Here we report functional studies of a testis-specific polycomb protein—SCML2. The Scml2 gene maps to the X chromosome. Intriguingly, the SCML2 protein localizes specifically to the XY chromatin in germ cells during male meiosis, which undergoes chromosome-wide transcriptional silencing. Disruption of Scml2 causes defects in spermatogenesis in mice. SCML2 associates with phosphorylated H2AX and a deubiquitinase, USP7. While localization of phosphorylated H2AX to the XY chromatin is SCML2-independent, USP7 localizes to the XY chromatin in an SCML2-dependent manner. Loss of SCML2 results in accumulation of H2A monoubiquitination in the XY chromatin in spermatocytes. These functional studies of SCML2 uncover a new molecular pathway that regulates H2A ubiquitination on the sex chromosomes during male meiosis.

Meiosis entry during spermatogenesis requires reprogramming from mitotic to meiotic gene expression profiles. Transcriptional regulation has been extensively studied in meiosis entry, but gain of function for master transcription factors is insufficient to down-regulate mitotic genes. RNA helicase YTHDC2 and its partner MEIOC emerge as essential posttranscriptional regulators of meiotic entry. However, it is unclear what governs the RNA binding specificity of YTHDC2/MEIOC. Here, we identified RNA binding protein RBM46 as a component of the YTHDC2/MEIOC complex. Testis-specific Rbm46 knockout in mice causes infertility with defective mitotic-to-meiotic transition, phenocopying global Ythdc2 or Meioc knockout. RBM46 binds to 3′ UTR of mitotic transcripts within 100 nucleotides from YTHDC2 U-rich motifs and targets these transcripts for degradation. Dysregulated RBM46 expression is associated with human male fertility disorders. These findings establish the RBM46/YTHDC2/MEIOC complex as the major posttranscriptional regulator responsible for down-regulating mitotic transcripts during meiosis entry in mammalian spermatogenesis, with implications for understanding meiosis-related fertility disorders.

Objective: To develop an imaging-derived biomarker for prediction of overall survival (OS) of pancreatic cancer by analyzing preoperative multiphase contrast-enhanced computed topography (CECT) using deep learning. Background: Exploiting prognostic biomarkers for guiding neoadjuvant and adjuvant treatment decisions may potentially improve outcomes in patients with resectable pancreatic cancer. Methods: This multicenter, retrospective study included 1516 patients with resected pancreatic ductal adenocarcinoma (PDAC) from 5 centers located in China. The discovery cohort (n=763), which included preoperative multiphase CECT scans and OS data from 2 centers, was used to construct a fully automated imaging-derived prognostic biomarker—DeepCT-PDAC—by training scalable deep segmentation and prognostic models (via self-learning) to comprehensively model the tumor-anatomy spatial relations and their appearance dynamics in multiphase CECT for OS prediction. The marker was independently tested using internal (n=574) and external validation cohorts (n=179, 3 centers) to evaluate its performance, robustness, and clinical usefulness. Results: Preoperatively, DeepCT-PDAC was the strongest predictor of OS in both internal and external validation cohorts [hazard ratio (HR) for high versus low risk 2.03, 95% confidence interval (CI): 1.50–2.75; HR: 2.47, CI: 1.35–4.53] in a multivariable analysis. Postoperatively, DeepCT-PDAC remained significant in both cohorts (HR: 2.49, CI: 1.89–3.28; HR: 2.15, CI: 1.14–4.05) after adjustment for potential confounders. For margin-negative patients, adjuvant chemoradiotherapy was associated with improved OS in the subgroup with DeepCT-PDAC low risk (HR: 0.35, CI: 0.19–0.64), but did not affect OS in the subgroup with high risk. Conclusions: Deep learning-based CT imaging-derived biomarker enabled the objective and unbiased OS prediction for patients with resectable PDAC. This marker is applicable across hospitals, imaging protocols, and treatments, and has the potential to tailor neoadjuvant and adjuvant treatments at the individual level.

Gallbladder carcinoma (GBC) is the most common malignancy of the bile duct and patients with GBC have extremely poor prognoses. PIK3CA, which encodes the phosphoinositide 3-kinase (PI3K) subunit p110α, is frequently mutated in many cancers, including GBC. The function of the E545K mutation in GBC is not fully understood. E545K mutation was determined in human GBC tissues by targeted sequencing. The effects of E545K mutation and PI3K selective inhibitor, A66 on GBC cells were evaluated using Cell Counting Kit-8 (CCK-8) cell Viability and transwell assays. The mechanisms of E545K mutation and A66 were analyzed by western blot and co-immunoprecipitation (Co-IP) assay. Subcutaneous xenograft models in nude mice were employed to evaluate the role of E545K mutation and A66 in GBC progression. The rate of PIK3CA E545K mutation in GBC patients was 6.15 %. And the survival of GBC patients was correlated with E545K mutation significantly (P < 0.05). The E545K mutation promoted proliferation, migration and invasion of GBC cells in vitro and tumor proliferation in vivo. A66 suppressed proliferation of GBC cells in vitro and tumor proliferation in vivo. The prognoses of patients with E545K mutation were worse than patients without this mutation. The E545K mutation promoted GBC progression through enhanced binding to EGFR and activating downstream akt activity. The PI3K selective inhibitor, A66, suppressed gallbladder carcinoma proliferation.