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Promyelocytic leukemia (PML) nuclear bodies (PML-NBs), a class of membrane-less cellular organelles, participate in various biological activities. PML-NBs are known as the core-shell-type nuclear body, harboring ‘client’ proteins in their core. Although multiple membrane-less organelles work in the oocyte nucleus, PML-NBs have been predicted to be absent from oocytes. Here, we show that some well-known PML clients (but not endogenous PML) co-localized with small ubiquitin-related modifier (SUMO) protein in the nucleolus and peri-centromeric heterochromatin of maturing oocytes. In oocytes devoid of PML-NBs, endogenous PML protein localized in the vicinity of chromatin. During and after meiotic resumption, PML co-localized with SUMO gathering around chromosomes. To examine the benefit of the PML-NB-free intranuclear milieu in oocytes, we deliberately assembled PML-NBs by microinjecting human PML-encoding plasmids into oocytes. Under conditions of limited SUMO availability, assembled PML-NBs tended to cluster. Upon proteotoxic stress, SUMO delocalized from peri-centromeric heterochromatin and co-localized with SC35 (a marker of nuclear speckles)-positive large compartments, which was disturbed by pre-assembled PML-NBs. These observations suggest that the PML-NB-free intranuclear environment helps reserve SUMO for emergent responses by redirecting the flux of SUMO otherwise needed to maintain PML-NB dynamics. more...

The functional roles of protein modification by small ubiquitin-like modifier (SUMO) proteins are not well understood compared to ubiquitination. Promyelocytic leukemia (PML) proteins are good substrates for SUMOylation, and PML-nuclear bodies (PML-NBs) may function as a platform for the PML SUMOylation. PML proteins are rapidly modified both with SUMO2/3 and SUMO1 after exposure to arsenite (As more...

BackgroundIdentification of female reproductive toxicants is currently based largely on integrated epidemiological and invivo toxicology data and, to a lesser degree, on mechanistic data. A uniform approach to systematically search, organize, integrate, and evaluate mechanistic evidence of female reproductive toxicity from various data types is lacking.ObjectiveWe sought to apply a key characteristics approach similar to that pioneered for carcinogen hazard identification to female reproductive toxicant hazard identification.MethodsA working group of international experts was convened to discuss mechanisms associated with chemical-induced female reproductive toxicity and identified 10 key characteristics of chemicals that cause female reproductive toxicity: 1) alters hormone receptor signaling; alters reproductive hormone production, secretion, or metabolism; 2) chemical or metabolite is genotoxic; 3) induces epigenetic alterations; 4) causes mitochondrial dysfunction; 5) induces oxidative stress; 6) alters immune function; 7) alters cell signal transduction; 8) alters direct cell–cell interactions; 9) alters survival, proliferation, cell death, or metabolic pathways; and 10) alters microtubules and associated structures. As proof of principle, cyclophosphamide and diethylstilbestrol (DES), for which both human and animal studies have demonstrated female reproductive toxicity, display at least 5 and 3 key characteristics, respectively. 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD), for which the epidemiological evidence is mixed, exhibits 5 key characteristics.DiscussionFuture efforts should focus on evaluating the proposed key characteristics against additional known and suspected female reproductive toxicants. Chemicals that exhibit one or more of the key characteristics could be prioritized for additional evaluation and testing. A key characteristics approach has the potential to integrate with pathway-based toxicity testing to improve prediction of female reproductive toxicity in chemicals and potentially prevent some toxicants from entering common use. https://doi.org/10.1289/EHP4971. more...

Inorganic arsenicals are well-known carcinogens, whereas arsenite (iAsIII) compounds are now recognized as potent therapeutic agents for several leukemias, and arsenic trioxide has been used for the treatment of recurrent acute promyelocytic leukemia (APL). However, recent clinical trials revealed that arsenite is not always effective for non-APL malignancies. Another arsenical, S-dimethylarsino-glutathione ([DMAIII(GS)]), which is a putative metabolic intermediate in the hepatic metabolism of iAsIII, shows promise for treating several types of lymphoma. However, the metabolism of [DMAIII(GS)] has not been well investigated, probably because [DMAIII(GS)] is not stable in biological fluids where the concentration of glutathione is low. In the present study, we injected [DMAIII(GS)] intravenously into mice and compared the tissue distribution and metabolic dynamics of [DMAIII(GS)] with those of sodium arsenite (NaAsO2). We found a unique organ preference for the distribution of [DMAIII(GS)] to the lung and brain in comparison to NaAsO2. Furthermore, [DMAIII(GS)] appeared to bind to serum albumin by exchanging its glutathione moiety quickly after administration, providing novel insights into the longer retention of [DMAIII(GS)] in plasma. more...

Oligo-astheno-teratozoospermia (OAT), a condition that includes low sperm number, low sperm motility and abnormal sperm morphology, is the commonest cause of male infertility. Because genetic analysis is frequently impeded by the infertility phenotype, the genetic basis of many of OAT conditions has been hard to verify. Here, we show that deficiency of ORP4, a sterol-binding protein in the oxysterol-binding protein (OSBP)-related protein family, causes male infertility due to severe OAT in mice. In ORP4-deficient mice, spermatogonia proliferation and subsequent meiosis occurred normally, but the morphology of elongating and elongated spermatids was severely distorted, with round-shaped head, curled back head or symplast. Spermatozoa derived from ORP4-deficient mice had little or no motility and no fertilizing ability in vitro. In ORP4-deficient testis, postmeiotic spermatids underwent extensive apoptosis, leading to a severely reduced number of spermatozoa. At the ultrastructural level, nascent acrosomes appeared to normally develop in round spermatids, but acrosomes were detached from the nucleus in elongating spermatids. These results suggest that ORP4 is essential for the postmeiotic differentiation of germ cells. more...

Particulate air pollution (PM 2.5) is a worldwide concern. Growing epidemiological evidence has shown pathophysiological effects of PM 2.5, not only on cardiovascular system but also on reproductive performance. The composition and physicochemical properties of PM 2.5 vary depending on the emission sources, climate conditions, and complex chemical reactions in the air. These factors make it difficult to understand the cause and mechanistic details of the adverse health effects of PM 2.5. Here, we show potential impacts of PM 2.5 on oocyte maturation in mice by utilizing diesel exhaust-derived secondary organic aerosol (SOA), a major component of urban PM 2.5. We found that the SOA destabilized microtubules of mouse oocytes and p-benzoquinone is one of the candidates for the microtubule-destabilizing compounds. We propose that some biologically reactive components of PM 2.5 should be prioritized for the regulation of atmospheric quality. more...

Arachidonic acid (AA) is remarkably enriched in phosphatidylinositol (PI). Studies using knockout mice of lysophosphatidylinositol acyltransferase 1, which selectively incorporates AA into PI, reveal that AA-containing PI plays a crucial role in cortical lamination and neuronal migration during brain development., Dietary arachidonic acid (AA) has roles in growth, neuronal development, and cognitive function in infants. AA is remarkably enriched in phosphatidylinositol (PI), an important constituent of biological membranes in mammals; however, the physiological significance of AA-containing PI remains unknown. In an RNA interference–based genetic screen using Caenorhabditis elegans, we recently cloned mboa-7 as an acyltransferase that selectively incorporates AA into PI. Here we show that lysophosphatidylinositol acyltransferase 1 (LPIAT1, also known as MBOAT7), the closest mammalian homologue, plays a crucial role in brain development in mice. Lpiat1−/− mice show almost no LPIAT activity with arachidonoyl-CoA as an acyl donor and show reduced AA contents in PI and PI phosphates. Lpiat1−/− mice die within a month and show atrophy of the cerebral cortex and hippocampus. Immunohistochemical analysis reveals disordered cortical lamination and delayed neuronal migration in the cortex of E18.5 Lpiat1−/− mice. LPIAT1 deficiency also causes disordered neuronal processes in the cortex and reduced neurite outgrowth in vitro. Taken together, these results demonstrate that AA-containing PI/PI phosphates play an important role in normal cortical lamination during brain development in mice. more...

SummaryMitochondria are dynamic organelles that change their morphology by active fusion and fission in response to cellular signaling and differentiation [1, 2]. The in vivo role of mitochondrial fission in mammals has been examined by using tissue-specific knockout (KO) mice of the mitochondria fission-regulating GTPase Drp1 [3, 4], as well as analyzing a human patient harboring a point mutation in Drp1 [5], showing that Drp1 is essential for embryonic and neonatal development and neuronal function. During oocyte maturation and aging, structures of various membrane organelles including mitochondria and the endoplasmic reticulum (ER) are changed dynamically [6, 7], and their organelle aggregation is related to germ cell formation and epigenetic regulation [8–10]. However, the underlying molecular mechanisms of organelle dynamics during the development and aging of oocytes have not been well understood. Here, we analyzed oocyte-specific mitochondrial fission factor Drp1-deficient mice and found that mitochondrial fission is essential for follicular maturation and ovulation in an age-dependent manner. Mitochondria were highly aggregated with other organelles, such as the ER and secretory vesicles, in KO oocyte, which resulted in impaired Ca2+ signaling, intercellular communication via secretion, and meiotic resumption. We further found that oocytes from aged mice displayed reduced Drp1-dependent mitochondrial fission and defective organelle morphogenesis, similar to Drp1 KO oocytes. On the basis of these findings, it appears that mitochondrial fission maintains the competency of oocytes via multiorganelle rearrangement. more...