Diversification of Ferredoxins across Living Organisms

Ferredoxins, iron-sulfur (Fe-S) cluster proteins, play a key role in oxidoreduction reactions. To date, evolutionary analysis of these proteins across the domains of life have been confined to observing the abundance of Fe-S cluster types (2Fe-2S, 3Fe-4S, 4Fe-4S, 7Fe-8S (3Fe-4s and 4Fe-4S) and 2[4Fe-4S]) and the diversity of ferredoxins within these cluster types was not studied. To address this research gap, here we propose a subtype classification and nomenclature for ferredoxins based on the characteristic spacing between the cysteine amino acids of the Fe-S binding motif as a subtype signature to assess the diversity of ferredoxins across the living organisms. To test this hypothesis, comparative analysis of ferredoxins between bacterial groups, Alphaproteobacteria and Firmicutes and ferredoxins collected from species of different domains of life that are reported in the literature has been carried out. Ferredoxins were found to be highly diverse within their types. Large numbers of alphaproteobacterial species ferredoxin subtypes were found in Firmicutes species and the same ferredoxin subtypes across the species of Bacteria, Archaea, and Eukarya, suggesting shared common ancestral origin of ferredoxins between Archaea and Bacteria and lateral gene transfer of ferredoxins from prokaryotes (Archaea/Bacteria) to eukaryotes. This study opened new vistas for further analysis of diversity of ferredoxins in living organisms.     

Phylogeography and genetic diversity of the commercially-collected Caribbean blue-legged hermit crab (Clibanarius tricolor)

Shifts from ornamental fish tanks to functional reef ecosystem aquaria has led to unprecedented pressure on marine invertebrate fisheries. The Caribbean blue-legged hermit crab (Clibanarius tricolor) is among the species targeted for functional reef tanks, valued for its role as an aquarium cleaner. Little is known about the biology of the species or the genetic landscape in which the increased collecting is happening. Here, we investigate the phylogeographic history and genetic diversity of C. tricolor through analysis of mitochondrial (CO1, 16S) and nuclear (H3) DNA. We test whether phylogeographic breaks for other invertebrates structure the genetic diversity of C. tricolor and explore additional factors that may govern structure, such as reproductive strategy, life history, habitat preference, adult mobility, demographic history, and vicariance events. Based on these three markers, we find high genetic diversity and connectivity and find no evidence to support the tested barriers as relevant to gene flow for C. tricolor. Rather, mitochondrial and nuclear markers infer high genetic diversity, panmixia, and demographic expansion during the Pleistocene. Our finding of panmixia makes it difficult to identify source or sink populations, but the absence of hierarchical structure inferred from mtDNA and nuDNA markers we use, high levels of genetic diversity and homogeneity for these same markers, and advantageous life history traits suggest C. tricolor is not currently at special risk; however, geographically restricted haplotypes and limitations within our study prevent us from making a strong conclusion about the sustainability of the fishery. Our work on the Caribbean blue-legged hermit crab highlights the importance of acquiring basic information on exploited species and reiterates that common regional forces may not equally impact connectivity among co-distributed species.

     

Host-cell DNA methylation patterns during high-risk HPV-induced carcinogenesis reveal a heterogeneous nature of cervical pre-cancer

Cervical cancer development following a persistent infection with high-risk human papillomavirus (hrHPV) is driven by additional host-cell changes, such as altered DNA methylation. In previous studies, we have identified 12 methylated host genes associated with cervical cancer and pre-cancer (CIN2/3). This study systematically analyzed the onset and DNA methylation pattern of these genes during hrHPV-induced carcinogenesis using a longitudinal in vitro model of hrHPV-transformed cell lines (n = 14) and hrHPV-positive cervical scrapings (n = 113) covering various stages of cervical carcinogenesis. DNA methylation analysis was performed by quantitative methylation-specific PCR (qMSP) and relative qMSP values were used to analyze the data. The majority of genes displayed a comparable DNA methylation pattern in both cell lines and clinical specimens. DNA methylation onset occurred at early or late immortal passage, and DNA methylation levels gradually increased towards tumorigenic cells. Subsequently, we defined a so-called cancer-like methylation-high pattern based on the DNA methylation levels observed in cervical scrapings from women with cervical cancer. This cancer-like methylation-high pattern was observed in 72% (38/53) of CIN3 and 55% (11/20) of CIN2, whereas it was virtually absent in hrHPV-positive controls (1/26). In conclusion, hrHPV-induced carcinogenesis is characterized by early onset of DNA methylation, typically occurring at the pre-tumorigenic stage and with highest DNA methylation levels at the cancer stage. Host-cell DNA methylation patterns in cervical scrapings from women with CIN2 and CIN3 are heterogeneous, with a subset displaying a cancer-like methylation-high pattern, suggestive for a higher cancer risk.     

GB virus C infection in Indonesian HIV‐positive patients

GB virus C (GBV‐C), a human virus of the Flaviviridae family that is structurally and epidemiologically closest to hepatitis C virus (HCV), has been reported to confer beneficial outcomes in HIV‐positive patients. However, the prevalence of GBV‐C in HIV‐positive individuals in Indonesia is unknown. Since GBV‐C is more prevalent in anti‐HCV positive patients than in anti‐HCV negative subjects, transmission of GBV‐C and HCV could be by the same method. This study examined the prevalence and molecular characteristics of GBV‐C infection in HIV patients in Yogyakarta, Indonesia. The prevalence of GBV‐C among HIV patients (n = 125, median age 31 years) based on the 5′UTR region was 111/125 (88.8%), including 39/48 (81.3%) and 72/77 (93.5%) HIV‐infected patients with and without HCV infection, respectively. GBV‐C isolates were of genotype 2a, 3 and 6 in 58.3%, 12.6% and 28.4% of patients, respectively. Patients with genotype 3 were significantly younger than those with genotypes 2a or 6 (P = 0.001 and P = 0.012, respectively). Genotypes 3 and 6 were significantly associated with injection drug use (P = 0.004 and P = 0.002, respectively) and HCV co‐infection (P < 0.001 for both genotypes), indicating a shared transmission route with HCV. In conclusion, the prevalence of GBV‐C among HIV‐positive patients in Indonesia is high, and three genotypes were detected, namely genotype 2a, 3 and 6.     

Online measurement of the viscosity in shake flasks enables monitoring of γ-PGA production in depolymerase knockout mutants of Bacillus subtilis with the phosphate-starvation inducible promoter Ppst

Poly-γ-glutamic acid (γ-PGA) is a biopolymer with a wide range of applications, mainly produced using Bacillus strains. The formation and concomitant secretion of γ-PGA increases the culture broth viscosity, while enzymatic depolymerisation and degradation of γ-PGA decreases the culture broth viscosity. In this study, the recently published ViMOS (V iscosity M onitoring O nline S ystem) is applied for optical online measurements of broth viscosity in eight parallel shake flasks. It is shown that the ViMOS is suitable to monitor γ-PGA production and degradation online in shake flasks. This online monitoring enables the detailed analysis of the P_pst promoter and γ-PGA depolymerase knockout mutants in genetically modified Bacillus subtilis 168. The P_pst promoter becomes active under phosphate starvation. The different single depolymerase knockout mutants are ∆ggt, ∆pgdS, ∆cwlO and a triple knockout mutant. An increase in γ-PGA yield in g_γ-PGA/g_glucose of 190% could be achieved with the triple knockout mutant compared to the P_pst reference strain. The single cwlO knockout also increased γ-PGA production, while the other single knockouts of ggt and pgdS showed no impact. Partial depolymerisation of γ-PGA occurred despite the triple knockout. The online measured data are confirmed with offline measurements. The online viscosity system directly reflects γ-PGA synthesis, γ-PGA depolymerisation, and changes in the molecular weight. Thus, the ViMOS has great potential to rapidly gain detailed and reliable information about new strains and cultivation conditions. The broadened knowledge will facilitate the further optimization of γ-PGA production.     

GPNMB plays a protective role against obesity-related metabolic disorders by reducing macrophage inflammatory capacity

Obesity is a global health problem that is often related to cardiovascular and metabolic diseases. Chronic low-grade inflammation in white adipose tissue (WAT) is a hallmark of obesity. Previously, during a search for differentially expressed genes in WAT of obese mice, we identified glycoprotein nonmetastatic melanoma protein B (GPNMB), of which expression was robustly induced in pathologically expanded WAT. Here, we investigated the role of GPNMB in obesity-related metabolic disorders utilizing GPNMB-deficient mice. When fed a high-fat diet (HFD), GPNMB-deficient mice showed body weight and adiposity similar to those of wild-type (WT) mice. Nonetheless, insulin and glucose tolerance tests revealed significant obesity-related metabolic disorders in GPNMB-KO mice compared with WT mice fed with HFD. Chronic WAT inflammation was remarkably worsened in HFD-fed GPNMB-KO mice, accompanied by a striking increase in crown-like structures, typical hallmarks for diseased WAT. Macrophages isolated from GPNMB-KO mice were observed to produce more inflammatory cytokines than those of WT mice, a difference abolished by supplementation with recombinant soluble GPNMB extracellular domain. We demonstrated that GPNMB reduced the inflammatory capacity of macrophages by inhibiting NF-κB signaling largely through binding to CD44. Finally, we showed that macrophage depletion by addition of clodronate liposomes abolished the worsened WAT inflammation and abrogated the exacerbation of metabolic disorders in GPNMB-deficient mice fed on HFD. Our data reveal that GPNMB negatively regulates macrophage inflammatory capacities and ameliorates the WAT inflammation in obesity; therefore we conclude that GPNMB is a promising therapeutic target for the treatment of metabolic disorders associated with obesity.