Mutations in TOP3A Cause a Bloom Syndrome-like Disorder

Bloom syndrome, caused by biallelic mutations in BLM, is characterized by prenatal-onset growth deficiency, short stature, an erythematous photosensitive malar rash, and increased cancer predisposition. Diagnostically, a hallmark feature is the presence of increased sister chromatid exchanges (SCEs) on cytogenetic testing. Here, we describe biallelic mutations in TOP3A in ten individuals with prenatal-onset growth restriction and microcephaly. TOP3A encodes topoisomerase III alpha (TopIIIα), which binds to BLM as part of the BTRR complex, and promotes dissolution of double Holliday junctions arising during homologous recombination. We also identify a homozygous truncating variant in RMI1, which encodes another component of the BTRR complex, in two individuals with microcephalic dwarfism. The TOP3A mutations substantially reduce cellular levels of TopIIIα, and consequently subjects’ cells demonstrate elevated rates of SCE. Unresolved DNA recombination and/or replication intermediates persist into mitosis, leading to chromosome segregation defects and genome instability that most likely explain the growth restriction seen in these subjects and in Bloom syndrome. Clinical features of mitochondrial dysfunction are evident in several individuals with biallelic TOP3A mutations, consistent with the recently reported additional function of TopIIIα in mitochondrial DNA decatenation. In summary, our findings establish TOP3A mutations as an additional cause of prenatal-onset short stature with increased cytogenetic SCEs and implicate the decatenation activity of the BTRR complex in their pathogenesis

In vitro anti-HIV activity of some Indian medicinal plant extracts

Background Human Immunodeficiency Virus (HIV) persists to be a significant public health issue worldwide. The current strategy for the treatment of HIV infection, Highly Active Antiretroviral Therapy (HAART), has reduced deaths from AIDS related disease, but it can be an expensive regime for the underdeveloped and developing countries where the supply of drugs is scarce and often not well tolerated, especially in persons undergoing long term treatment. The present therapy also has limitations of development of multidrug resistance, thus there is a need for the discovery of novel anti-HIV compounds from plants as a potential alternative in combating HIV disease. Methods Ten Indian medicinal plants were tested for entry and replication inhibition against laboratory adapted strains HIV-1IIIB, HIV-1Ada5 and primary isolates HIV-1UG070, HIV-1VB59 in TZM-bl cell lines and primary isolates HIV-1UG070, HIV-1VB59 in PM1 cell lines. The plant extracts were further evaluated for toxicity in HEC-1A epithelial cell lines by transwell epithelial model. Results The methanolic extracts of Achyranthes aspera, Rosa centifolia and aqueous extract of Ficus benghalensis inhibited laboratory adapted HIV-1 strains (IC80 3.6–118 μg/ml) and primary isolates (IC80 4.8–156 μg/ml) in TZM-bl cells. Methanolic extract of Strychnos potatorum, aqueous extract of Ficus infectoria and hydroalcoholic extract of Annona squamosa inhibited laboratory adapted HIV-1 strains (IC80 4.24–125 μg/ml) and primary isolates (IC80 18–156 μg/ml) in TZM-bl cells. Methanolic extracts of Achyranthes aspera and Rosa centifolia, (IC801-9 μg/ml) further significantly inhibited HIV-1 primary isolates in PM1cells. Methanolic extracts of Tridax procumbens, Mallotus philippinensis, Annona reticulate, aqueous extract of Ficus benghalensis and hydroalcoholic extract of Albizzia lebbeck did not exhibit anti-HIV activity in all the tested strains. Methanolic extract of Rosa centifolia also demonstrated to be non-toxic to HEC-1A epithelial cells and maintained epithelial integrity (at 500 μg/ml) when tested in transwell dual-chamber. Conclusion These active methanolic extracts of Achyranthes aspera and Rosa centifolia, could be further subjected to chemical analysis to investigate the active moiety responsible for the anti-HIV activity. Methanolic extract of Rosa centifolia was found to be well tolerated maintaining the epithelial integrity of HEC-1A cells in vitro and thus has potential for investigating it further as candidate microbicide

Mycobacterium tuberculosis UvrD1 and UvrD2 helicases unwind G-quadruplex DNA

Unresolved G-quadruplex (G4) DNA secondary structures impede DNA replication and can lead to DNA breaks and to genome instability. Helicases are known to unwind G4 structures and thereby facilitate genome duplication. Escherichia coli UvrD is a multifunctional helicase that participates in DNA repair, recombination and replication. Previously, we had demonstrated a novel role of E. coli UvrD helicase in resolving G4 structures. Mycobacterium tuberculosis genome encodes two orthologs of E. coli UvrD helicase, UvrD1 and UvrD2. It is unclear whether UvrD1 or UvrD2 or both helicases unwind G4 DNA structures. Here, we demonstrate that M. tuberculosis UvrD1 and UvrD2 unwind G4 tetraplexes. Both helicases were proficient in resolving previously characterized tetramolecular G4 structures in an ATP hydrolysis and single-stranded 3 `-tail-dependent manner. Notably, M. tuberculosis UvrD1 and UvrD2 were efficient in unwinding G4 structures derived from the potential G4 forming sequences present in the M. tuberculosis genome. These data suggest an extended role for M. tuberculosis UvrD1 and UvrD2 helicases in resolving G4 DNA structures and provide insights into the maintenance of genome integrity via G4 DNA resolution

Escherichia coli and Neisseria gonorrhoeae UvrD helicase unwinds G4 DNA structures

G-quadruplex (G4) secondary structures have been implicated in various biological processes, including gene expression, DNA replication and telomere maintenance. However, unresolved G4 structures impede replication progression which can lead to the generation of DNA double-strand breaks and genome instability. Helicases have been shown to resolve G4 structures to facilitate faithful duplication of the genome. Escherichia coli UvrD (EcUvrD) helicase plays a crucial role in nucleotide excision repair, mismatch repair and in the regulation of homologous recombination. Here, we demonstrate a novel role of E. coli and Neisseria gonorrhoeae UvrD in resolving G4 tetraplexes. EcUvrD and N. gonorrhoeae UvrD were proficient in unwinding previously characterized tetramolecular G4 structures. Notably, EcUvrD was equally efficient in resolving tetramolecular and bimolecular G4 DNA that were derived from the potential G4-forming sequences from the genome of E. coli. Interestingly, in addition to resolving intermolecular G4 structures, EcUvrD was robust in unwinding intramolecular G4 structures. These data for the first time provide evidence for the role of UvrD in the resolution of G4 structures, which has implications for the in vivo role of UvrD helicase in G4 DNA resolution and genome maintenance

Outcome of Cataract Surgery among Eye Camp Patients of Eastern Nepal

Background: Outreach camp is helpful for reducing the prevalence of blindness due to cataracts and refractive error. The objective was to evaluate the visual outcome of cataract surgery among camp patients operated at a high volume center of Eastern Nepal.  Materials and Methods: Descriptive cross sectional study design was used. The comprehensive eye screening program was conducted and those having cataracts were referred to base hospital for cataract surgery, patients were reexamined at the base hospital again and a small incision cataract surgery with posterior chamber intraocular lens was implanted at Biratnagar Eye Hospital, Biratnagar and Sagarmatha Choudhary Eye Hospital, Lahan. The pre-operative vision was taken during screening and post-operative vision was taken after one-month by performing refraction by trained personnel. Data were extracted from the hospital record; analysis was done using SPSS version 17.0. Descriptive analysis by using frequency, parent age and association was done by using Chi square test with a P-value <0.05 considered as significant.  Results: A total of 1426 camp patients were operated. Among them 1113 (78%) were visited after one month post operative follow up at the campsite. There were 744 (52%) female and bilateral affected eyes were 735(52%). Pre-operative presenting visual acuity was <3/60 (37%), which was reduced to (0.2%) after one month follows up of cataract surgery. There was a significant association in pre-operative VA with aided one month to follow up VA.  Conclusion: The outcome of the cataract surgery of camp patients operated at base hospitals was good in high-volume hospitals. Follow up after one month was good practice to provide services and measure the outcome of cataract surgery

Mycobacterium tuberculosis DinG Is a Structure-specific Helicase That Unwinds G4 DNA IMPLICATIONS FOR TARGETING G4 DNA AS A NOVEL THERAPEUTIC APPROACH

The significance of G-quadruplexes and the helicases that resolve G4 structures in prokaryotes is poorly understood. The Mycobacterium tuberculosis genome is GC-rich and contains >10,000 sequences that have the potential to form G4 structures. In Escherichia coli, RecQ helicase unwinds G4 structures. However, RecQ is absent in M. tuberculosis, and the helicase that participates in G4 resolution in M. tuberculosis is obscure. Here, we show that M. tuberculosis DinG (MtDinG) exhibits high affinity for ssDNA and ssDNA translocation with a 5' -> 3' polarity. Interestingly, MtDinG unwinds overhangs, flap structures, and forked duplexes but fails to unwind linear duplex DNA. Our data with DNase I footprinting provide mechanistic insights and suggest that MtDinG is a 5' -> 3' polarity helicase. Notably, in contrast to E. coli DinG, MtDinG catalyzes unwinding of replication fork and Holliday junction structures. Strikingly, we find that MtDinG resolves intermolecular G4 structures. These data suggest that MtDinG is a multifunctional structure-specific helicase that unwinds model structures of DNA replication, repair, and recombination as well as G4 structures. We finally demonstrate that promoter sequences of M. tuberculosis PE_PGRS2, mce1R, and moeB1 genes contain G4 structures, implying that G4 structures may regulate gene expression in M. tuberculosis. We discuss these data and implicate targeting G4 structures and DinG helicase in M. tuberculosis could be a novel therapeutic strategy for culminating the infection with this pathogen

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Not AvailableThe evaluation of soil quality is essential in monitoring the long term effects of rice cultivation. Present study investigated the effects of long term rice cultivation on soil properties and organic C pools and identified indicators for monitoring soil quality in Ghaggar-flood plains of hot arid India. Soil samples were collected from fields with 0, 10, 20, 30 and 40 years of rice cultivation. The study revealed that electrical conductivity (EC) and exchangeable sodium percentage (ESP) increased after 30–40 years of rice cultivation. Available nutrients increased with increas- ing years of rice cultivation. The organic carbon pools namely, total organic carbon (TOC), Walkley Black carbon (WBC) and particulate organic carbon (POC) were increased above 50% in 20 and above years of rice cultivation. The TOC and POC were increased by 40.6 to 132.4% and 31.7% to 104.8% in 10 to 40 years of rice cultivation. Cation exchange capacity, WBC, ESP and CaCO3 could serve as soil monitoring indicators of long term rice cultivation in arid region. The findings clearly indicated that long term rice cultivation could aggravate soil salinity and have negative impact on soil quality in arid environment.Not Availabl