The dazzling array of basal branches in the mtDNA macrohaplogroup M from India as inferred from complete genomes

Many efforts based on complete mitochondrial DNA (mtDNA) genomeshave been made to depict the global mtDNA landscape, but thephylogeny of Indian macrohaplogroup M has not yet been resolvedin detail. To fill this lacuna, we took the same strategy asin our recent analysis of Indian mtDNA macrohaplogroup N andselected 56 mtDNAs from over 1,200 samples across India forcomplete sequencing, with the intention to cover all Indianautochthonous M lineages. As a result, the phylogenetic statusof previously identified haplogroups based on control-regionand/or partial coding-region information, such as M2, M3, M4,M5, M6, M30, and M33, was solidified or redefined here. Moreover,seven novel basal M haplogroups (viz., M34–M40) were identified,and yet another five singular branches of the M phylogeny werediscovered in the present study. The comparison of matrilinealcomponents among India, East Asia, Southeast Asia, and Oceaniaat the deepest level yielded a star-like and nonoverlappingpattern, reflecting a rapid mode of modern human dispersal alongthe Asian coast after the initial "out-of-Africa" event.     

Fine structure of the organ of attachment of the teleost, Garra gotyla gotyla (Ham)

We describe the morphology of the attachment organ (AO) of the teleost, Garra gotyla gotyla (Cyprinidae). It is located ventrally around the mouth opening and used by the species for attachment to submerged rocks in sub-Himalayan streams and rivers where it lives. The AO consists of three crescentic parts and a central callus part. Scanning electron microscopy (SEM) shows the former to possess numerous tubercles, each of which bears about 23–27 curved spines. Light microscopy shows the epidermis of the tuberculated parts to possess one type of cell arranged into 7–8 rows. Transmission electron microscopy (TEM) reveals these cells to contain abundant tonofilaments (hence called the filament cells). The epidermis of the callus part possesses the filament cells and additionally mucous cells, which are absent in the tuberculated parts. The superficial epidermis is apparently keratinized (thickness: 5–8 μm), and a part of the cells of the outer row is modified into spines. These cells show a thick plasma membrane envelope and possess mucous granules (diameter: 0.1–0.3 μm) and bundles of tonofilaments. The cells of the inner two to four rows possess similar organelles and additionally, prominent Golgi bodies and rough endoplasmic reticulum. Immunohistochemically, the cells of the outer row and the spines stain positively for cytokeratin. The cells of the innermost rows (five to eight) possess few tonofilaments and no mucous granules. It is evident that the filament cells of the mid- to upper epidermis are specialized for the production of mucous granules and tonofilaments, which is unique for the teleost epidermis concerned. It appears that the tuberculated parts with spines assist in anchorage and interlocking with the substratum, while the central callus part probably utilizes both suction and frictional mechanisms, and mucous secretion protects the spines from damage during anchorage and abrasion.     

Establishment and characterization of a buffalo (Bubalus bubalis) mammary epithelial cell line

Background

The objective of this study was to establish the buffalo mammary epithelial cell line (BuMEC) and characterize its mammary specific functions.

Methodology

Buffalo mammary tissue collected from the slaughter house was processed enzymatically to obtain a heterogenous population of cells containing both epithelial and fibroblasts cells. Epithelial cells were purified by selective trypsinization and were grown in a plastic substratum. The purified mammary epithelial cells (MECs) after several passages were characterized for mammary specific functions by immunocytochemistry, RT-PCR and western blot.

Principal Findings

The established buffalo mammary epithelial cell line (BuMEC) exhibited epithelial cell characteristics by immunostaining positively with cytokeratin 18 and negatively with vimentin. The BuMEC maintained the characteristics of its functional differentiation by expression of β-casein, κ-casein, butyrophilin and lactoferrin. BuMEC had normal growth properties and maintained diploid chromosome number (2n = 50) before and after cryopreservation. A spontaneously immortalized buffalo mammary epithelial cell line was established after 20 passages and was continuously subcultured for more than 60 passages without senescence.

Conclusions

We have established a buffalo mammary epithelial cell line that can be used as a model system for studying mammary gland functions.     

BRCA1-mediated signaling pathways in ovarian carcinogenesis

The link between loss or defect in functional BRCA1 and predisposition for development of ovarian and breast cancer is well established. Germ-line mutations in BRCA1 are responsible for both hereditary breast and ovarian cancer, which is around 5–10% for all breast and 10–15% of all ovarian cancer cases. However, majority of cases of ovarian cancer are sporadic in nature. The inactivation of cellular BRCA1 due to mutations or loss of heterozygosity is one of the most commonly observed events in such cases. Complement-resistant retroviral BRCA1 vector, MFG-BRCA1, is the only approved gene therapy for ovarian cancer patients by the Federal and Drug Administration. Given the limited available information, there is a need to evaluate the effects of BRCA1 on the global gene expression pattern for better understanding the etiology of the disease. Here, we use Ingenuity Pathway Knowledge Base to examine the differential pattern of global gene expression due to stable expression of BRCA1 in the ovarian cancer cell line, SKOV3. The functional analysis detected at least five major pathways that were significantly (p < 0.05) altered. These include: cell to cell signaling and interaction, cellular function and maintenance, cellular growth and proliferation, cell cycle and DNA replication, and recombination repair. In addition, we were able to detect several biologically relevant genes that are central for various signaling networks involved in cellular homeostasis; TGF-β1, TP53, c-MYC, NF-κB and TNF-α. This report provides a comprehensive rationale for tumor suppressor function(s) of BRCA1 in ovarian carcinogenesis.     

Increased risk of lung cancer associated with a functionally impaired polymorphic variant of the human DNA glycosylase NEIL2

Human NEIL2, one of five oxidized base-specific DNA glycosylases, is unique in preferentially repairing oxidative damage in transcribed genes. Here we show that depletion of NEIL2 causes a 6-7-fold increase in spontaneous mutation frequency in the HPRT gene of the V79 Chinese hamster lung cell line. This prompted us to screen for NEIL2 variants in lung cancer patients' genomic DNA. We identified several polymorphic variants, among which R103Q and R257L were frequently observed in lung cancer patients. We then characterized these variants biochemically, and observed a modest decrease in DNA glycosylase activity relative to the wild type (WT) only with the R257L mutant protein. However, in reconstituted repair assays containing WT NEIL2 or its R257L and R103Q variants together with other DNA base excision repair (BER) proteins (PNKP, Polβ, Lig IIIα and XRCC1) or using NEIL2-FLAG immunocomplexes, an ~5-fold decrease in repair was observed with the R257L variant compared to WT or R103Q NEIL2, apparently due to the R257L mutant's lower affinity for other repair proteins, particularly Polβ. Notably, increased endogenous DNA damage was observed in NEIL2 variant (R257L)-expressing cells relative to WT cells. Taken together, our results suggest that the decreased DNA repair capacity of the R257L variant can induce mutations that lead to lung cancer development.     

Targeting tumors with peptides from natural sources

Peptide-based therapies offer the potential for non-genotoxic, genotype-specific alternatives, or adjuvants, to the current range of traditional cancer treatments. Such a patient-tailored cancer-cell-directed therapeutic approach should have fewer side effects and could well be more effective than the current drug- or combination-based regimens. Here, we review the potential of novel natural anticancer peptides such as necrotic peptides, apoptotic peptides, function-blocking peptides, antiangiogenic peptides and immunostimulatory peptides in the context of their ability to induce tumor regression. We focus on the therapeutic prospects of anticancer peptides and their possible application in tumor therapy.     

Decolorization and purification of crude protease from Rhizopus oryzae by activated charcoal and its electrophoretic analysis

Activated charcoal decolorized and partially purified the protease from a crude extract of solid state fermentation of wheat bran by Rhizopus oryzae. Treatment for 5 min was sufficient. Depending on the initial colour intensity of crude, the charcoal to crude extract ratio could be optimized to achieve 90% decolorization, 85% enzyme recovery, and over a 3-fold purification, even up to 20-fold variation in batch size (from 1 ml to 20 ml crude extract). Decolorization followed the Freundlich and the Langmuir models, the Freundlich constant, n, being 2.74. Partial purification was confirmed by native PAGE and the protease band identified by gelatin-PAGE. SDS-PAGE showed the protease consisted of two sub-units (about 22 and 24 kDa). List of symbols: c _o, initial solute concentration in liquid before adsorption; c ^*, equilibrium solute concentration in liquid after adsorption; k, empirical constant for Freundlich adsorption isotherm; U, unit of protease activity; v, volume of solution per unit weight of adsorbent.     

Structural characterization of NRAS isoform 5

It was recently discovered that the NRAS isoform 5 (20 amino acids) is expressed in melanoma and results in a more aggressive cell phenotype. This novel isoform is responsible for increased phosphorylation of downstream targets such as AKT, MEK, and ERK as well as increased cellular proliferation. This structure report describes the NMR solution structure of NRAS isoform 5 to be used as a starting point to understand its biophysical interactions. The isoform is highly flexible in aqueous solution, but forms a helix‐turn‐coil structure in the presence of trifluoroethanol as determined by NMR and CD spectroscopy.