The mammary gland microenvironment during postlactational involution shares similarities with inflammation, including high matrix metalloproteinase activity, fibrillar collagen deposition, and release of bioactive fragments of fibronectin and laminin. Because inflammation can promote tumorigenesis, we evaluated whether the tissue microenvironment of the involuting gland is also promotional. Extracellular matrix was isolated from mammary glands of nulliparous rats or rats with mammary glands undergoing weaning-induced involution. Using these matrices as substratum, nulliparous matrix was found to promote ductal organization of normal mammary epithelial MCF-12A cells in three-dimensional culture and to suppress invasion of mammary tumor MDA-MB-231 cells in transwell filter assays. Conversely, involution matrix failed to support ductal development in normal cells and promoted invasiveness in tumor cells. To evaluate the effects of these matrices on metastasis in vivo, MDA-MB-231 cells, premixed with Matrigel, nulliparous matrix, or involution matrix, were injected into mammary fat pads of nude mice. Metastases to lung, liver, and kidney were increased in the involution matrix group, and correlated with a twofold increase in tumor vascular endothelial growth factor expression and increased angiogenesis. These data suggest that the mammary gland microenvironment becomes promotional for tumor cell dissemination during involution, thus providing a plausible mechanism to explain the high rate of metastases that occur with pregnancy-associated breast cancer.

Mitochondrial mRNAs in kinetoplastids require extensive U-insertion/deletion editing that progresses 3 ′ -to-5 ′ in small blocks, each directed by a guide RNA (gRNA), and exhibits substrate and developmental stage-specificity by unsolved mechanisms. Here, we address compositionally related factors, collectively known as the mitochondrial RNA-binding complex 1 (MRB1) or gRNAbinding complex (GRBC), that contain gRNA, have a dynamic protein composition, and transiently associate with several mitochondrial factors including RNA editing core complexes (RECC) and ribosomes. MRB1 controls editing by still unknown mechanisms. We performed the first next-generation sequencing study of native subcomplexes of MRB1, immunoselected via either RNA helicase 2 (REH2), that binds RNA and associates with unwinding activity, or MRB3010, that affects an early editing step. The particles contain either REH2 or MRB3010 but share the core GAP1 and other proteins detected by RNA photo-crosslinking. Analyses of the first editing blocks indicate an enrichment of several initiating gRNAs in the MRB3010-purified complex. Our data also indicate fast evolution of mRNA 3 ′ ends and strain-specific alternative 3 ′ editing within 3 ′ UTR or C-terminal protein-coding sequence that could impact mitochondrial physiology. Moreover, we found robust specific copurification of edited and pre-edited mRNAs, suggesting that these particles may bind both mRNA and gRNA editing substrates. We propose that multiple subcomplexes of MRB1 with different RNA/protein composition serve as a scaffold for specific assembly of editing substrates and RECC, thereby forming the editing holoenzyme. The MRB3010-subcomplex may promote early editing through its preferential recruitment of initiating gRNAs.

Translational control during cell division determines when cells start a new cell cycle, how fast they complete it, the number of successive divisions, and how cells coordinate proliferation with available nutrients. The translational efficiencies of mRNAs in cells progressing synchronously through the mitotic cell cycle, while preserving the coupling of cell division with cell growth, remain uninvestigated. We now report comprehensive ribosome profiling of a yeast cell size series from the time of cell birth, to identify mRNAs under periodic translational control. The data reveal coordinate translational activation of mRNAs encoding lipogenic enzymes late in the cell cycle including Acc1p, the rate-limiting enzyme acetyl-CoA carboxylase. An upstream open reading frame (uORF) confers the translational control of and adjusts Acc1p protein levels in different nutrients. The uORF is relevant for cell division because its ablation delays cell cycle progression, reduces cell size, and suppresses the replicative longevity of cells lacking the Sch9p protein kinase regulator of ribosome biogenesis. These findings establish an unexpected relationship between lipogenesis and protein synthesis in mitotic cell divisions.

The short splice variant of the basic helix-loop-helix Per-Arnt-Sim transcription factor Singleminded-2, SIM2s, has been implicated in development and is frequently lost or reduced in primary breast tumors. Here, we show that loss of Sim2s causes aberrant mouse mammary gland ductal development with features suggestive of malignant transformation, including increased proliferation, loss of polarity, down-regulation of Ecadherin, and invasion of the surrounding stroma. Additionally, knockdown of SIM2s in MCF-7 breast cancer cells contributed to an epithelial-mesenchymal transition (EMT) and increased tumorigenesis. In both Sim2؊/؊ mammary glands and SIM2s-depleted MCF7 cells, these changes were associated with increased SLUG and MMP2 levels. SIM2s protein was detectable on the SLUG promoter, and overexpression of SIM2s repressed expression from a SLUG-controlled reporter in a dose-dependent manner. To our knowledge, SIM2s is the first protein shown to bind and repress the SLUG promoter, providing a plausible explanation for the development role and breast tumor-suppressive activity of SIM2s. Together, our results suggest that SIM2s is a key regulator of mammary-ductal development and that loss of SIM2s expression is associated with an invasive, EMT-like phenotype.

Dynamic evolutionary processes and complex structure make the Y chromosome among the most diverse and least understood regions in mammalian genomes. Here, we present an annotated assembly of the male specific region of the horse Y chromosome (eMSY), representing the first comprehensive Y assembly in odd-toed ungulates. The eMSY comprises single-copy, equine specific multi-copy, PAR transposed, and novel ampliconic sequence classes. The eMSY gene density approaches that of autosomes with the highest number of retained X–Y gametologs recorded in eutherians, in addition to novel Y-born and transposed genes. Horse, donkey and mule testis RNAseq reveals several candidate genes for stallion fertility. A novel testis-expressed XY ampliconic sequence class, ETSTY7, is shared with the parasite Parascaris genome, providing evidence for eukaryotic horizontal transfer and inter-chromosomal mobility. Our study highlights the dynamic nature of the Y and provides a reference sequence for improved understanding of equine male development and fertility.