Necessity of HuR/ELAVL1 for the activation-induced cytidine deaminase-dependent decrease in topoisomerase 1 in antibody diversification.

Activation-induced cytidine deaminase (AID)-dependent DNA cleavage is the initial event of antibody gene-diversification processes such as class switch recombination (CSR) and somatic hypermutation (SHM). We previously reported the requirement of an AID-dependent decrease of topoisomerase 1 (Top1) for efficient DNA cleavage, but the underlying molecular mechanism has remained elusive. This study focuses on HuR/ELAVL1, a protein that binds to AU-rich elements in RNA. HuR-knockout (KO) CH12 cells derived from murine B lymphoma cells were found to have lower CSR and hypermutation efficiencies due to decreased AID-dependent DNA cleavage levels. The HuR-KO CH12 cells do not show impairment in cell cycles and Myc expression, which have been reported in HuR-reduced spleen B cells. Furthermore, drugs that scavenge reactive oxygen species (ROS) do not rescue the lower CSR in HuR-KO CH12 cells, meaning that ROS or decreased c-Myc protein amount is not the reason for the deficiencies of CSR and hypermutation in HuR-KO CH12 cells. We show that HuR binds to Top1 mRNA and that complete deletion of HuR abolishes AID-dependent repression of Top1 protein synthesis in CH12 cells. Additionally, reduction of CSR to IgG3 in HuR-KO cells is rescued by knockdown of Top1, indicating that elimination of the AID-dependent Top1 decrease is the cause of the inefficiency of DNA cleavage, CSR and hypermutation in HuR-KO cells. These results show that HuR is required for initiation of antibody diversification and acquired immunity through the regulation of AID-dependent DNA cleavage by repressing Top1 protein synthesis.

Green synthesis of zinc oxide nanoparticles using Elaeagnus angustifolia L. leaf extracts and their multiple in vitro biological applications.

Due to their versatile applications, ZnONPs have been formulated by several approaches, including green chemistry methods. In the current study, convenient and economically viable ZnONPs were produced using Elaeagnus angustifolia (EA) leaf extracts. The phytochemicals from E. angustifolia L. are believed to serve as a non-toxic source of reducing and stabilizing agents. The physical and chemical properties of ZnONPs were investigated employing varying analytical techniques (UV, XRD, FT-IR, EDX, SEM, TEM, DLS and Raman). Strong UV-Vis absorption at 399 nm was observed for green ZnONPs. TEM, SEM and XRD analyses determined the nanoscale size, morphology and crystalline structure of ZnONPs, respectively. The ZnONPs were substantiated by evaluation using HepG2 (IC50: 21.7 µg mL-1) and HUH7 (IC50: 29.8 µg mL-1) cancer cell lines and displayed potential anticancer activities. The MTT cytotoxicity assay was conducted using Leishmania tropica "KWH23" (promastigotes: IC50, 24.9 µg mL-1; and amastigotes: IC50, 32.83 µg mL-1). ZnONPs exhibited excellent antimicrobial potencies against five different bacterial and fungal species via the disc-diffusion method, and their MIC values were calculated. ZnONPs were found to be biocompatible using human erythrocytes and macrophages. Free radical scavenging tests revealed excellent antioxidant activities. Enzyme inhibition assays were performed and revealed excellent potential. These findings suggested that EA@ZnONPs have potential applications and could be used as a promising candidate for clinical development.

Biallelic Missense Mutation in the ECEL1 Underlies Distal Arthrogryposis Type 5 (DA5D).

Distal arthrogryposis (DA) is a heterogeneous sub-group of arthrogryposis multiplex congenita (AMC), mostly characterized by having congenital contractures affecting hands, wrists, feet, and ankles. Distal arthrogryposis is mostly autosomal dominantly inherited, while only one sub-type DA type 5D is inherited in an autosomal recessive manner. Clinically, DA5D is described having knee extension contractures, micrognathia, distal joint contractures, clubfoot, ptosis, contractures (shoulders, elbows, and wrists), and scoliosis. Using whole exome sequencing (WES) followed by Sanger sequencing, we report on a first familial case of DA5D from Pakistani population having a novel biallelic missense mutation (c.158C>A, p.Pro53Leu) in the ECEL1 gene. Our result support that homozygous mutations in ECEL1 causes DA5D and expands the clinical and allelic spectrum of ECEL1 related contracture syndromes.