Defective Maturation Research Articles

Rare homozygous mutation in TUBB8 associated with oocyte maturation defect-2 in a consanguineous mating family

PurposeVariations in many genes may lead to the occurrence of oocyte maturation defects. To investigate the genetic basis of oocyte maturation defects, we performed clinical and genetic analysis of a pedigree.MethodsThe proband with oocyte maturation defect-2 receiving ovulation induction therapy and her parents were selected for clinical detection, whole exome sequencing and Sanger sequencing. One unrelated healthy woman received ovulation induction therapy as control. Mutations were assessed after frequency screening of public exome databases. Then homozygous variants shared by the proband and her parents were selected.ResultsArrest of oocytes maturation was observed. A new missense mutation in TUBB8 (TUBB8: NM_177,987: exon 2: c. C161T: p. A54V) was identified, which was shown to be rare compared with public databases. The variant was highly conserved among primates, and was suggested to be deleterious by online software prediction.ConclusionsThe homozygote of this variant (TUBB8: NM_ 177,987: exon 2:c.C161T: p.A54V) might affect spindle assembly, cause arrest of oocyte maturation and lead to oocyte maturation defect-2.

Neonatal therapy with clenbuterol and salmeterol restores spinogenesis and dendritic complexity in the dentate gyrus of the Ts65Dn model of Down syndrome

Down syndrome (DS), a neurodevelopmental disorder caused by triplication of chromosome 21, is characterized by intellectual disability. In DS, defective neurogenesis causes an overall reduction in the number of neurons populating the brain and defective neuron maturation causes dendritic hypotrophy and reduction in the density of dendritic spines. No effective therapy currently exists for the improvement of brain development in individuals with DS. Drug repurposing is a strategy for identifying new medical use for approved drugs. A drug screening campaign showed that the β2-adrenergic receptor (β2-AR) agonists clenbuterol hydrochloride (CLEN) and salmeterol xinafoate (SALM) increase the proliferation rate of neural progenitor cells from the Ts65Dn model of DS. The goal of the current study was to establish their efficacy in vivo, in the Ts65Dn model. We found that, at variance with the in vitro experiments, treatment with CLEN or SALM did not restore neurogenesis in the hippocampus of Ts65Dn mice treated during the postnatal (P) period P3-P15. In Ts65Dn mice treated with CLEN or SALM, however, dendritic spine density and dendritic arborization of the hippocampal granule cells were restored and the lowest dose tested here (0.01 mg/kg/day) was sufficient to elicit these effects. CLEN and SALM are used in children as therapy for asthma and, importantly, they pass the blood-brain barrier. Our study suggests that treatment with these β2-AR agonists may be a therapy of choice in order to correct dendritic development in DS but is not suitable to rescue neurogenesis.

Cardiomyocyte Maturation: New Phase in Development.

Maturation is the last phase of heart development that prepares the organ for strong, efficient, and persistent pumping throughout the mammal's lifespan. This process is characterized by structural, gene expression, metabolic, and functional specializations in cardiomyocytes as the heart transits from fetal to adult states. Cardiomyocyte maturation gained increased attention recently due to the maturation defects in pluripotent stem cell-derived cardiomyocyte, its antagonistic effect on myocardial regeneration, and its potential contribution to cardiac disease. Here, we review the major hallmarks of ventricular cardiomyocyte maturation and summarize key regulatory mechanisms that promote and coordinate these cellular events. With advances in the technical platforms used for cardiomyocyte maturation research, we expect significant progress in the future that will deepen our understanding of this process and lead to better maturation of pluripotent stem cell-derived cardiomyocyte and novel therapeutic strategies for heart disease.

A Metformin-Responsive Metabolic Pathway Controls Distinct Steps in Gastric Progenitor Fate Decisions and Maturation.

Cellular metabolism plays important functions in dictating stem cell behaviors, although its role in stomach epithelial homeostasis has not been evaluated in depth. Here, we show that the energy sensor AMP kinase (AMPK) governs gastric epithelial progenitor differentiation. Administering the AMPK activator metformin decreases epithelial progenitor proliferation and increases acid-secreting parietal cells (PCs) in mice and organoids. AMPK activation targets Krüppel-like factor 4 (KLF4), known to govern progenitor proliferation and PC fate choice, and PGC1α, which we show controls PC maturation after their specification. PC-specific deletion of AMPKα or PGC1α causes defective PC maturation, which could not be rescued by metformin. However, metformin treatment still increases KLF4 levels and suppresses progenitor proliferation. Thus, AMPK activates KLF4 in progenitors to reduce self-renewal and promote PC fate, whereas AMPK-PGC1α activation within the PC lineage promotes maturation, providing a potential suggestion for why metformin increases acid secretion and reduces gastric cancer risk in humans.

Identification of RNA Exosome Cofactors in Neuronal Cells to Probe Disease Mechanism

The RNA exosome, a 10‐subunit complex that mediates both RNA processing and degradation, plays a critical role in defining cellular expression profiles. This complex is ubiquitously expressed, essential, and critical for fundamental cellular functions, such as ribosomal RNA processing. Recent studies have linked mutations in genes encoding multiple subunits of the complex to tissue‐specific human disease. For example, missense mutations in the human EXOSC3 gene, which encodes an RNA exosome subunit, cause Pontocerebellar Hypoplasia type 1b (PCH1b), a disease characterized by atrophy of the pons and cerebellum. The missense mutations encode single amino acid changes in conserved regions of the EXOSC3 protein. How these amino acid substitutions confer tissue‐specific phenotypes is not known. One possible mechanism underlying the distinct disease phenotypes could be a decrease in the interaction of the RNA exosome complex with cofactors that confer specificity for RNA targets. However, most studies that identify and characterize RNA exosome cofactors have been carried out in budding yeast and thus, do not provide insight into whether tissue‐specific cofactors could exist. Our studies use immunoprecipitation from neuronal cell culture (N2A) and relevant mouse tissues to define RNA exosome cofactors. Biochemical experiments that employ cultured N2A cells were used to identify RNA exosome‐interacting proteins in both the nucleus and the cytoplasm. Preliminary results from this analysis reveal an association between the RNA exosome and a large complex of cytoplasmic tRNA ligase enzymes, which could link defects in tRNA maturation to disease pathology. We are extending these studies to explore the possibility of tissue‐specific cofactors by immunoprecipitating EXOSC3 and analyzing co‐purifying proteins from the mouse cerebellum (affected in disease) and cortex (unaffected). To complement these approaches to identify tissue‐specific RNA exosome cofactors, we are also exploring whether amino acid changes that are linked to disease alter cofactor interactions. These studies, which are being performed in N2A cells, have identified cofactors that show altered interactions with the EXOSC3 variants present in disease. These studies will provide insight into both the functional consequences of amino acid substitutions in the RNA exosome that cause disease and the role of cofactors in conferring RNA target specificity.

RNA‐Binding Protein QKI is a critical pre‐RNA splicing regulator for cardiac development and function

RNA‐binding protein QKI (also known as Quaking) belongs to hnRNP K‐homology (KH)‐domain protein family and is implicated a novel regulator in pre‐RNA splicing. As previously shown, the QKI homolog how potentially regulates heart lumen formation and function in fruit fly. However, QKI’s function in mammalian heart development is largely unknown. We found that Qki specifically expressed in mouse developing hearts as early as E7.0 and this high‐level expression was maintained in the hearts to adulthood. Consistent with this finding, by using human embryonic stem cells (hESCs)‐cardiomyocyte differentiation system, QKI expression was found up‐regulated at Day 6, a critical time frame that cardiogenic progenitor transition into early cardiomyocytes. To determine the biological function of QKI in cardiogenesis, we generated hESCsQKIdef by using CRISPR/Cas9 gene editing technology and analyzed the physiological impact of QKI‐deficiency to cardiomyocyte differentiation and maturation. As expected, we did not find defect in pluripotency in hESCsQKIdef, but the contractile function was dramatically reduced in hESCsQKIdef‐derived cardiomyocytes. Based on scRNA‐seq and clustering analyses, there was no significant difference in the distribution of the clusters when compared hESCsQKIdef‐cardiomyocyte group to normal control group, suggesting that the cardiomyocyte differentiation was largely unaffected in hESCsQKIdef. However, scRNA‐seq demonstrated a handful of genes encoding sarcomere proteins were significantly altered in cardiomyocyte clusters, which included ACTN2, ACTA1, MYH7, and MYL2, etc., indicating a significant defect in cardiomyocyte maturation. Bulk RNA‐seq data not only further confirmed this notion, but also provided additional depth for detailed analysis on the QKI‐mediated pre‐RNA splicing events. In conjunction with QKI‐binding motif search and RNA immunoprecipitation (RIP) analysis, we were able to demonstrate that QKI was a key pre‐RNA splicing regulator for sets of genes involved in cardiac myofibrillogenesis and cardiac excitation‐contraction (E‐C) coupling, which ultimately impacted on cardiomyocyte maturation and function. Additional analysis of cardiac developmental defects in Qkibgeo mutant mice further validated that QKI was an indispensable splicing regulator in cardiovascular development. Our results not only revealed a novel mechanism by which QKI regulates pre‐mRNA splicing in cardiogenesis, but also implied that QKI was likely involved in the pathogenesis of certain forms of congenital heart defects and cardiomyopathies.Support or Funding InformationThis work was supported in part by National Institute of Health P01HL134599

Homozygous Splice Site Mutation in ZP1 Causes Familial Oocyte Maturation Defect.

In vitro fertilization (IVF) involves controlled ovarian hyperstimulation using hormones to produce large numbers of oocytes. The success of IVF is tightly linked to the availability of mature oocytes. In most cases, about 70% to 80% of the oocytes are mature at the time of retrieval, however, in rare instances, all of them may be immature, implying that they were not able to reach the metaphase II (MII) stage. The failure to obtain any mature oocytes, despite a well conducted ovarian stimulation in repeated cycles is a very rare cause of primary female infertility, for which the underlying suspected genetic factors are still largely unknown. In this study, we present the whole exome sequencing analysis of a consanguineous Turkish family comprising three sisters with a recurrent oocyte maturation defect. Analysis of the data reveals a homozygous splice site mutation (c.1775-3C>A) in the zona pellucida glycoprotein 1 (ZP1) gene. Minigene experiments show that the mutation causes the retention of the intron 11 sequence between exon 11 and exon 12, resulting in a frameshift and the likely production of a truncated protein.

Functional and Pharmacological Characterization of the Rare CFTR Mutation W361R.

Understanding the functional consequence of rare cystic fibrosis (CF) mutations is mandatory for the adoption of precision therapeutic approaches for CF. Here we studied the effect of the very rare CF mutation, W361R, on CFTR processing and function. We applied western blot, patch clamp and pharmacological modulators of CFTR to study the maturation and ion transport properties of pEGFP-WT and mutant CFTR constructs, W361R, F508del and L69H-CFTR, expressed in HEK293 cells. Structural analyses were also performed to study the molecular environment of the W361 residue. Western blot showed that W361R-CFTR was not efficiently processed to a mature band C, similar to F508del CFTR, but unlike F508del CFTR, it did exhibit significant transport activity at the cell surface in response to cAMP agonists. Importantly, W361R-CFTR also responded well to CFTR modulators: its maturation defect was efficiently corrected by VX-809 treatment and its channel activity further potentiated by VX-770. Based on these results, we postulate that W361R is a novel class-2 CF mutation that causes abnormal protein maturation which can be corrected by VX-809, and additionally potentiated by VX-770, two FDA-approved small molecules. At the structural level, W361 is located within a class-2 CF mutation hotspot that includes other mutations that induce variable disease severity. Analysis of the 3D structure of CFTR within a lipid environment indicated that W361, together with other mutations located in this hotspot, is at the edge of a groove which stably accommodates lipid acyl chains. We suggest this lipid environment impacts CFTR folding, maturation and response to CFTR modulators.

Altered Tregs Differentiation and Impaired Autophagy Correlate to Atherosclerotic Disease.

Atherosclerosis is a progressive vascular disease representing the primary cause of morbidity and mortality in developed countries. Formerly, atherosclerosis was considered as a mere passive accumulation of lipids in blood vessels. However, it is now clear that atherosclerosis is a complex and multifactorial disease, in which the involvement of immune cells and inflammation play a key role. A variety of studies have shown that autophagy—a cellular catalytic mechanism able to remove injured cytoplasmic components in response to cellular stress—may be proatherogenic. So far, in this context, its role has been investigated in smooth muscle cells, macrophages, and endothelial cells, while the function of this catabolic protective process in lymphocyte functionality has been overlooked. The few studies carried out so far, however, suggested that autophagy modulation in lymphocyte subsets may be functionally related to plaque formation and development. Therefore, in this research, we aimed at better clarifying the role of lymphocyte subsets, mainly regulatory T cells (Tregs), in human atherosclerotic plaques and in animal models of atherosclerosis investigating the contribution of autophagy on immune cell homeostasis. Here, we investigate basal autophagy in a mouse model of atherosclerosis, apolipoprotein E (ApoE)-knockout (KO) mice, and we analyze the role of autophagy in driving Tregs polarization. We observed defective maturation of Tregs from ApoE-KO mice in response to tumor growth factor-β (TGFβ). TGFβ is a well-known autophagy inducer, and Tregs maturation defects in ApoE-KO mice seem to be related to autophagy impairment. In this work, we propose that autophagy underlies Tregs maturation, advocating that the study of this process in atherosclerosis may open new therapeutic strategies.

USP16 counteracts mono-ubiquitination of RPS27a and promotes maturation of the 40S ribosomal subunit.

Establishment of translational competence represents a decisive cytoplasmic step in the biogenesis of 40S ribosomal subunits. This involves final 18S rRNA processing and release of residual biogenesis factors, including the protein kinase RIOK1. To identify novel proteins promoting the final maturation of human 40S subunits, we characterized pre-ribosomal subunits trapped on RIOK1 by mass spectrometry, and identified the deubiquitinase USP16 among the captured factors. We demonstrate that USP16 constitutes a component of late cytoplasmic pre-40S subunits that promotes the removal of ubiquitin from an internal lysine of ribosomal protein RPS27a/eS31. USP16 deletion leads to late 40S subunit maturation defects, manifesting in incomplete processing of 18S rRNA and retarded recycling of late-acting ribosome biogenesis factors, revealing an unexpected contribution of USP16 to the ultimate step of 40S synthesis. Finally, ubiquitination of RPS27a appears to depend on active translation, pointing at a potential connection between 40S maturation and protein synthesis.

48 Burn Injury Impedes Late Erythropoiesis via Decreased Alpha Hemoglobin Stabilizing Protein and siglec1

Abstract Introduction Transfusions are of no benefit to burn patients with chronic anemia. We are elucidating the mechanisms of erythropoietin resistant anemia in burn injury using a mouse model in addition to a contrived system with human PBMCs. We have found that Alpha Hemoglobin Stabilizing Protein (AHSP), an erythroblast specific protein essential to effective hemoglobinization, is decreased in PBMC derived erythroblasts (EB) from burn patients (male + female). Hemoglobinization begins at the polychromatic EB (PolyE) stage and intensifies at the orthochromatic EB (OrthoE) stage, culminating in ejection of the nucleus and the cell becoming a reticulocyte (Reti). EB needs the support of a central island macrophage (EBI MØ) until they progress to maturation. We have seen in bone marrow from mice (male) following burn injury that Siglec1, an essential adhesion molecule is decreased in EBI MØ. It is these MØ that provide iron for the assembly of the hemoglobin unit in late erythropoiesis. EB maturation defects seen in burn injury could be due to the intrinsic changes in EB phenotype or the phenotype of supporting EBI MØ or a combination of the two. Our goal was to investigate in the animal model using male and female mice, how myeloid/erythroid intrinsic changes after burn impact the progression of late erythropoiesis from an EB and EBI MØ perspectives. Methods With IACUC approval, 12 male and female mice were anesthetized and subjected to sham burn or scald burn (15% TBSA) upon dorsal immersion in a 100oC water bath for 8 seconds. Animals were sacrificed on post burn day 7 (PBD 7). Total bone marrow cells (TBM) were harvested from bilateral femurs. Cells were characterized using CD71, Ter119, and Syto16 antibodies. EBI MØ were identified using a combination of F4/80, ERHR3, Vcam1, Ly6G and Siglec 1 antibodies. Geometric mean of fluorescence intensity (MFI) and cell counts were obtained by flow cytometric analysis and presented as x103 per million TBM cells. Peripheral blood was harvested via cardiac puncture and hemoglobin concentrations (g/dL) were obtained using a hematology analyzer. Results Among the erythroblasts, EEBs were 2.4 fold higher with a 2.4 fold lower AHSP expression in PBD 7 mice compared to sham as shown in left, top and bottom figures. Similarly, more EEBs were associated with EBIMØ with a lower Siglec1 expression as shown in right, top and bottom figures. Blood Hbg concentrations were lower in burn (sham=15.5±0.7 vs PBD 7= 13.1±0.5; p< 0.01). Conclusions The combined influence of Siglec1 and AHSP expression interferes with effective hemoglobinization impeding late maturation of erythroblasts following burn injury. Whether there is a synergistic action between the two deficiencies is being studied. Applicability of Research to Practice Underpinning the mechanism of erythropoiesis will lead to better options to ineffective transfusions in burn patients with anemia.

Microfluidic modeling of bone marrow pathophysiology

Human bone-marrow-on-a-chip improves drug studies of hematopoietic cell function and identifies a neutrophil maturation defect in a genetic disorder.

GORAB promotes embryonic lung maturation through antagonizing AKT phosphorylation, versican expression, and mesenchymal cell migration.

Embryonic development of the alveolar sac of the lung is dependent upon multiple signaling pathways to coordinate cell growth, migration, and the formation of the extracellular matrix. Here, we identify GORAB as a regulator of embryonic alveolar sac formation as genetically disrupting the Gorab gene in mice resulted in fatal saccular maturation defects characterized by a thickened lung mesenchyme. This abnormality is not associated with impairments in cellular proliferation and death, but aberrantly increased protein kinase B (AKT) phosphorylation, elevated Vcan transcription, and enhanced migration of mesenchymal fibroblasts. Genetically augmenting PDGFRα, a potent activator of AKT in lung mesenchymal cells, recapitulated the alveolar phenotypes, whereas disrupting PDGFRα partially rescued alveolar phenotypes in Gorab-deficient mice. Overexpressing or suppressing Vcan in primary embryonic lung fibroblasts could, respectively, mimic or attenuate alveolar sac-like phenotypes in a co-culture model. These findings suggest a role of GORAB in negatively regulating AKT phosphorylation, the expression of Vcan, and the migration of lung mesenchyme fibroblasts, and suggest that alveolar sac formation resembles a patterning event that is orchestrated by molecular signaling and the extracellular matrix in the mesenchyme.

Diethylstilbestrol exposure disrupts mouse oocyte meiotic maturation in vitro through affecting spindle assembly and chromosome alignment

An adverse tendency induced by the environmental estrogens in female reproductive health is one serious problem worldwide. Diethylstilbestrol (DES), as a synthetic estrogen, is still used as an animal growth stimulant in terrestrial livestock and aquaculture illegally. It has been reported to negatively affect ovarian function and oogenesis. Nevertheless, the mechanism and toxicity of DES on oocyte meiotic maturation are largely unknown. Herein, we found that DES (40 μM) intervened in mouse oocyte maturation and first polar body extrusion (PBE) was decreased in vitro. Cell cycle analysis showed meiotic process was disturbed with oocytes arrested at metaphase I (MI) stage after DES exposure. Further study showed that DES exposure disrupted the spindle assembly and chromosome alignment, which then continuously provoke the spindle assemble checkpoint (SAC). We also observed that the acetylation levels of α-tubulin were dramatically increased in DES-treated oocytes. In addition, the dynamics of actin were also affected. Moreover, the distribution patterns of estrogen receptor α (ERα) were altered in DES-treated oocyte, as indicated by the significant signals accumulation in the spindle area. However, ERα inhibitor failed to rescue the defects of oocyte maturation caused by DES. Of note, the same phenomenon was observed in estrogen-treated oocytes. Collectively, we showed that DES exposure lead to the oocyte meiotic failure via impairing the spindle assembly and chromosome alignment. Our research is helpful to understand how environmental estrogen affects female germ cells and contribute to design the potential therapies to preserve fertility especially for occupational exposure.

The green tea polyphenol epigallocatechin-3-gallate (EGCG) restores CDKL5-dependent synaptic defects in vitro and in vivo

CDKL5 deficiency disorder (CDD) is a rare X-linked neurodevelopmental disorder that is characterised by early-onset seizures, intellectual disability, gross motor impairment, and autistic-like features. CDD is caused by mutations in the cyclin-dependent kinase-like 5 (CDKL5) gene that encodes a serine/threonine kinase with a predominant expression in the brain. Loss of CDKL5 causes neurodevelopmental alterations in vitro and in vivo, including defective dendritic arborisation and spine maturation, which most likely underlie the cognitive defects and autistic features present in humans and mice.Here, we show that treatment with epigallatocathechin-3-gallate (EGCG), the major polyphenol of green tea, can restore defects in dendritic and synaptic development of primary Cdkl5 knockout (KO) neurons. Furthermore, defective synaptic maturation in the hippocampi and cortices of adult Cdkl5-KO mice can be rescued through the intraperitoneal administration of EGCG, which is however not sufficient to normalise behavioural CDKL5-dependent deficits. EGCG is a pleiotropic compound with numerous cellular targets, including the dual-specificity tyrosine-phosphorylation-regulated kinase 1A (DYRK1A) that is selectively inhibited by EGCG. DYRK1A controls dendritic development and spine formation and its deregulation has been implicated in neurodevelopmental and degenerative diseases. Treatment with another DYRK1A inhibitor, harmine, was capable of correcting neuronal CDKL5-dependent defects; moreover, DYRK1A levels were upregulated in primary Cdkl5-KO neurons in concomitance with increased phosphorylation of Tau, a well-accepted DYRK1A substrate. Altogether, our results indicate that DYRK1A deregulation may contribute, at least in part, to the neurodevelopmental alterations caused by CDKL5 deficiency.

Casein Kinase 1δ Stabilizes Mature Axons by Inhibiting Transcription Termination of Ankyrin.

After axon outgrowth and synapse formation, the nervous system transitions to a stable architecture. In C.elegans, this transition is marked by the appearance of casein kinase 1δ (CK1δ) in the nucleus. In CK1δ mutants, neurons continue to sprout growth cones into adulthood, leading to a highly ramified nervous system. Nervous system architecture in these mutants is completely restored by suppressor mutations in ten genes involved in transcription termination. CK1δ prevents termination by phosphorylating and inhibiting SSUP-72. SSUP-72 would normally remodel the C-terminal domain of RNA polymerase in anticipation of termination. The antitermination activity of CK1δ establishes the mature state of a neuron by promoting the expression of the long isoform of a single gene, the cytoskeleton protein Ankyrin.

Sperm DNA Fragmentation: A New Guideline for Clinicians.

Sperm DNA integrity is crucial for fertilization and development of healthy offspring. The spermatozoon undergoes extensive molecular remodeling of its nucleus during later phases of spermatogenesis, which imparts compaction and protects the genetic content. Testicular (defective maturation and abortive apoptosis) and post-testicular (oxidative stress) mechanisms are implicated in the etiology of sperm DNA fragmentation (SDF), which affects both natural and assisted reproduction. Several clinical and environmental factors are known to negatively impact sperm DNA integrity. An increasing number of reports emphasizes the direct relationship between sperm DNA damage and male infertility. Currently, several assays are available to assess sperm DNA damage, however, routine assessment of SDF in clinical practice is not recommended by professional organizations. This article provides an overview of SDF types, origin and comparative analysis of various SDF assays while primarily focusing on the clinical indications of SDF testing. Importantly, we report four clinical cases where SDF testing had played a significant role in improving fertility outcome. In light of these clinical case reports and recent scientific evidence, this review provides expert recommendations on SDF testing and examines the advantages and drawbacks of the clinical utility of SDF testing using Strength-Weaknesses-Opportunities-Threats (SWOT) analysis.

Endo-Lysosomal Cation Channels and Infectious Diseases.

Among the infectious diseases caused by pathogenic microorganisms such as bacteria, viruses, parasites, or fungi, the most prevalent ones today are malaria, tuberculosis, influenza, HIV/AIDS, Ebola, dengue fever, and methicillin-resistant Staphylococcus aureus (MRSA) infection, and most recently Covid-19 (SARS-CoV2). Others with a rather devastating history and high fatality rates such as plague, cholera, or typhus seem less threatening today but have not been eradicated, and with a declining efficacy of current antibiotics they ought to be watched carefully. Another emerging issue in this context is health-care associated infection. About 100,000 hospitalized patients in the USA ( www.cdc.gov ) and 33,000 in Europe ( https://www.ecdc.europa.eu ) die each year as a direct consequence of an infection caused by bacteria resistant to antibiotics. Among viral infections, influenza is responsible for about 3-5 million cases of severe illness, and about 250,000 to 500,000 deaths annually ( www.who.int ). About 37 million people are currently living with HIV infection and about one million die from it each year. Coronaviruses such as MERS-CoV, SARS-CoV, but in particular the recent outbreak of Covid-19 (caused by SARS-CoV2) have resulted in large numbers of infections worldwide with an estimated several hundred thousand deaths (anticipated fatality rate: <5%). With a comparatively low mortality rate dengue virus causes between 50 and 100 million infections every year, leading to 50,000 deaths. In contrast, Ebola virus is the causative agent for one of the deadliest viral diseases. The Ebola outbreak in West Africa in 2014 is considered the largest outbreak in history with more than 11,000 deaths. Many of the deadliest pathogens such as Ebola virus, influenza virus, mycobacterium tuberculosis, dengue virus, and cholera exploit the endo-lysosomal trafficking system of host cells for penetration into the cytosol and replication. Defects in endo-lysosomal maturation, trafficking, fusion, or pH homeostasis can efficiently reduce the cytotoxicity caused by these pathogens. Most of these functions critically depend on endo-lysosomal membrane proteins such as transporters and ion channels. In particular, cation channels such as the mucolipins (TRPMLs) or the two-pore channels (TPCs) are involved in all of these aspects of endo-lysosomal integrity. In this review we will discuss the correlations between pathogen toxicity and endo-lysosomal cation channel function, and their potential as drug targets for infectious disease therapy.

In vitro strategies to enhance oocyte developmental competence.

Oocyte quality influences early embryonic survival, establishment and maintenance of pregnancy, fetal development and adult diseases. The developmental competence of oocytes is acquired gradually and increases with follicular development. The ability of an oocyte to develop into an embryo depends on, having enough specific information in the form of mRNA or proteins. If this information is insufficient, defects in nuclear or cytoplasmic maturation, or in both processes, may arise and thus affect the in vitro development of fertilized oocytes. The greater developmental competence of oocytes aspirated from larger follicles is reported as compared with smaller follicles. Oocyte developmental competence is greatly correlated with the morphology of the cumulus oocyte complexes (COCs). Apart from morphological or biochemical markers, molecular markers have also been investigated. Until now, no specific markers of oocyte developmental competence could be described for the oocyte developmental competence. To, utilize female germplasm to its maximum, there is a need to enhance developmental competence of lesser competent oocytes derived from the follicles which are not fully grown. The oocyte pre-maturation and maturation conditions affect gene expression not only in the oocyte but till the blastocyst stages too. Strategies have been discussed in this review would be useful to enhance the developmental competence of oocytes.

Genome regulation by long noncoding RNAs in neonatal heart maturation and congenital heart defects

Genome regulation by long noncoding RNAs in neonatal heart maturation and congenital heart defects