Effects of cesium on in vitro myoblast differentiation: An electron microscopic study

Summary This paper describes the microscopic evidence supporting a cesium-induced delay in the fusion of chick embryo myoblast membranes during in vitro myogenic differentiation. We have recently demonstrated that the sharp decrease in the conductivity and permittivity of the membranes of these myogenic cells at the time of fusion is delayed 30 h by the addition of cesium to the culture medium (Santini et al., Biochim. Biophys. Acta 945:56–64; 1988). We report here that this delay in fusion is substantiated by direct microscopic observation and that cesium also induces ultrastructural changes in the myoblast cells themselves. Possible mechanisms by which cesium may cause both the delay in fusion as well as the ultrastructural changes observed are discussed. This investigation was partially supported by an Italian Consiglio Nazionale delle Ricerche grant 85.00.304.02 (to P. L. I.).     

Genetic differentiation and detection of cryptic species in the genus Lepismachilis (Insecta: Microcoryphia) from the Western Mediterranean region

Different species of the bristletail genus Lepismachilis were collected in 14 localities in Italy and Spain and an allozyme electrophoretic survey was carried out to estimate the degree of genetic variability and differentiation at intra- and interspecific levels. Four morphological species were initially identified (L osellai, L. y-signata, L. affinis, L. targionii), but the electrophoretic analysis demonstrated the presence of two additional species among the individuals of L. targionii (Lepismachilis spl and sp2). The validity of these species and their differentiation from L targionii were demonstrated by the fixation of alternative allelic patterns at several loci (7 in Lepismachilis spl and 8 in Lepismachilis sp2), coupled with fixed, previously undetected, morphological differences. In addition, Lepismachilis sp2 was sympatric with L. targionii in three collecting sites, where the fixation of alternative allelic patterns unequivocally demonstrated reproductive isolation. Genetic variability did not seem to be correlated with local ecological factors, and differences between species should rather be explained by different historical factors. Low levels of gene flow, estimated with two different indirect methods, were observed in L. targionii and L. y-signata, and were due to high levels of structuring among populations. Genetic differentiation among conspecific populations was not correlated to their geographical arrangement and the presence of loci fixed for different alleles among them suggested that stochastic factors (such as genetic drift) may have played a role in determining genetic differentiation of geographically isolated populations. Genetic divergence values indicated that the six species are well differentiated and allozyme profiles were diagnostic for all of them. On the other hand, allozyme data did not provide adequate information to resolve evolutionary relationships among the species, nor did they confirm the validity of the two subgenera (Lepismachilis and Berlesilis) in which the genus Lepismachilis is traditionally divided.     

Phylogenetic island disequilibrium: evidence for ongoing long‐term population dynamics in two Mediterranean butterflies

Aim Our aims were to verify the existence of phylogenetic disequilibrium between butterfly lineages at the subcontinental scale for islands and the nearest mainland and to test the capacity of islands for hosting ancestral populations of butterflies and the significance of such relict populations. Location The western Mediterranean continental area of Europe and North Africa together with several large and small islands (Balearics, Tuscan Archipelago, Aeolian Archipelago, Capri, Sardinia, Sicily, Corsica). Methods Using geometric morphometrics, the shape of male genitalia was analysed in two common butterflies (Pyronia cecilia and Pyronia tithonus), whose spatial heterogeneity in the Mediterranean region has recently been described. Observed patterns in genital shapes were compared with shapes predicted for islands and fossil islands to assess the contribution of historical and current events in accounting for the transition from a refugial model to an equilibrium model. Measurements were taken for 473 specimens in 90 insular and mainland sites. Results The shape of the genitalia of populations of most islands differed substantially from that predicted by the equilibrium hypothesis while closely fitting the refugial hypothesis. The comparison between different models strongly suggests that islands maintain ancestral lineages similar to those living in Spain (P. cecilia) and France (P. tithonus). A high correlation between observed and predicted patterns on islands and fossil islands occurs during the first steps of modelled introgressive hybridization while the following steps exposed a successively lower fit, suggesting that the process from a refugial to an equilibrium situation is highly skewed towards an earlier non‐equilibrium. Main conclusions The observed non‐equilibrium pattern supports the refugial hypothesis, suggesting that an ancestral lineage was originally distributed from Spain to Italy, and also occupied offshore islands. This lineage, replaced in Italy, has persisted on the islands owing to their isolation. A comparison of the distribution patterns for genetic and morphometric markers in several species indicates that the situation highlighted for Pyronia may represent a common biogeographic feature for many Mediterranean butterflies.     

Position and sequence conservation in Amniota of polymorphic enhancer HS1.2 within the palindrome of IgH 3'Regulatory Region

Background  

The Immunoglobulin heavy chain (IgH) 3' Regulatory Region (3'RR), located at the 3' of the constant alpha gene, plays a crucial role in immunoglobulin production. In humans, there are 2 copies of the 3'RR, each composed of 4 main elements: 3 enhancers and a 20 bp tandem repeat. The single mouse 3'RR differs from the two human ones for the presence of 4 more regulative elements with the double copy of one enhancer at the border of a palindromic region.     

Modeling HIV quasispecies evolutionary dynamics

Background

During the HIV infection several quasispecies of the virus arise, which are able to use different coreceptors, in particular the CCR5 and CXCR4 coreceptors (R5 and X4 phenotypes, respectively). The switch in coreceptor usage has been correlated with a faster progression of the disease to the AIDS phase. As several pharmaceutical companies are starting large phase III trials for R5 and X4 drugs, models are needed to predict the co-evolutionary and competitive dynamics of virus strains.

Results

We present a model of HIV early infection which describes the dynamics of R5 quasispecies and a model of HIV late infection which describes the R5 to X4 switch. We report the following findings: after superinfection (multiple infections at different times) or coinfection (simultaneous infection by different strains), quasispecies dynamics has time scales of several months and becomes even slower at low number of CD4+ T cells. Phylogenetic inference of chemokine receptors suggests that viral mutational pathway may generate a large variety of R5 variants able to interact with chemokine receptors different from CXCR4. The decrease of CD4+ T cells, during AIDS late stage, can be described taking into account the X4-related Tumor Necrosis Factor dynamics.

Conclusion

The results of this study bridge the gap between the within-patient and the inter-patients (i.e. world-wide) evolutionary processes during HIV infection and may represent a framework relevant for modeling vaccination and therapy.

     

ZMAT3 hypomethylation contributes to early senescence of preadipocytes from healthy first‐degree relatives of type 2 diabetics

Senescence of adipose precursor cells (APC) impairs adipogenesis, contributes to the age‐related subcutaneous adipose tissue (SAT) dysfunction, and increases risk of type 2 diabetes (T2D). First‐degree relatives of T2D individuals (FDR) feature restricted adipogenesis, reflecting the detrimental effects of APC senescence earlier in life and rendering FDR more vulnerable to T2D. Epigenetics may contribute to these abnormalities but the underlying mechanisms remain unclear. In previous methylome comparison in APC from FDR and individuals with no diabetes familiarity (CTRL), ZMAT3 emerged as one of the top‐ranked senescence‐related genes featuring hypomethylation in FDR and associated with T2D risk. Here, we investigated whether and how DNA methylation changes at ZMAT3 promote early APC senescence. APC from FDR individuals revealed increases in multiple senescence markers compared to CTRL. Senescence in these cells was accompanied by ZMAT3 hypomethylation, which caused ZMAT3 upregulation. Demethylation at this gene in CTRL APC led to increased ZMAT3 expression and premature senescence, which were reverted by ZMAT3 siRNA. Furthermore, ZMAT3 overexpression in APC determined senescence and activation of the p53/p21 pathway, as observed in FDR APC. Adipogenesis was also inhibited in ZMAT3‐overexpressing APC. In FDR APC, rescue of ZMAT3 methylation through senolytic exposure simultaneously downregulated ZMAT3 expression and improved adipogenesis. Interestingly, in human SAT, aging and T2D were associated with significantly increased expression of both ZMAT3 and the P53 senescence marker. Thus, DNA hypomethylation causes ZMAT3 upregulation in FDR APC accompanied by acquisition of the senescence phenotype and impaired adipogenesis, which may contribute to FDR predisposition for T2D.     

ER-lysosome lipid transfer protein VPS13C/PARK23 prevents aberrant mtDNA-dependent STING signaling

Mutations in VPS13C cause early-onset, autosomal recessive Parkinson’s disease (PD). We have established that VPS13C encodes a lipid transfer protein localized to contact sites between the ER and late endosomes/lysosomes. In the current study, we demonstrate that depleting VPS13C in HeLa cells causes an accumulation of lysosomes with an altered lipid profile, including an accumulation of di-22:6-BMP, a biomarker of the PD-associated leucine-rich repeat kinase 2 (LRRK2) G2019S mutation. In addition, the DNA-sensing cGAS-STING pathway, which was recently implicated in PD pathogenesis, is activated in these cells. This activation results from a combination of elevated mitochondrial DNA in the cytosol and a defect in the degradation of activated STING, a lysosome-dependent process. These results suggest a link between ER-lysosome lipid transfer and innate immune activation in a model human cell line and place VPS13C in pathways relevant to PD pathogenesis.