Complement-Fixation Test for Detection of Herpes-like Viruses in Cell Cultures of Burkitt''s Lymphoma

Although biopsies of Burkitt's tumors contained no detectable complement-fixing (CF) antigen or antigens, tumor cell lines contained CF antigen or antigens related to the presence of a herpes-like virus particle.     

RFLP and RAPD mapping of the rice gm2 gene that confers resistance to biotype 1 of gall midge (Orseolia oryzae)

Gm2 is dominant gene conferring resistance to biotype 1 of gall midge (Orseolia oryzae Wood-Mason), the major dipteran pest of rice. The gene was mapped by restriction fragment length polymorphism (RFLP) analysis of a set of 40 recombinant inbred lines derived from a cross between the resistant variety Phalguna and the susceptible landrace ARC 6650. The gene is located on chromosome 4 at a position 1.3 cM from marker RG329 and 3.4 cM from RG476. Since the low (28%) polymorphism of this indica x indica cross hindered full coverage of the genome with RFLP markers, the mapping was checked by random amplified polymorphic DNA (RAPD)/bulked segregant analysis. Through the use of 160 RAPD primers, the number of polymorphic markers was increased from 43 to 231. Two RAPD primers amplified loci that co-segregated with resistance/susceptibility. RFLP mapping of these loci showed that they are located 0.7 cM and 2.0 cM from RG476, confirming the location of Gm2 in this region of chromosome 4. Use of these DNA markers will accelerate breeding for gall midge resistance by permitting selection of the Gm2 gene independently of the availability of the insect.     

Lipid content of brain, brain membrane lipid domains, and neurons from acid sphingomyelinase deficient mice

The cholesterol, sphingolipid, and glycerophospholipid content of total brain, of detergent-resistant membranes prepared from the total brain, and of cerebellar granule cells differentiated in culture from wild type (WT) and acid sphingomyelinase knockout (ASMKO) were studied. Brains derived from 7-month-old ASMKO animals showed a fivefold higher level of sphingomyelin and a significant increase in ganglioside content, mainly because of monosialogangliosides GM3 and GM2 accumulation, while the cholesterol and glycerophospholipid content was unchanged with respect to WT animals. An increase in sphingomyelin, but not in gangliosides, was also detected in cultured cerebellar granule neurons from ASMKO mice, indicating that ganglioside accumulation is not a direct consequence of the enzyme defect. When a detergent-resistant membrane fraction was prepared from ASMKO brains, we observed that a higher detergent-to-protein ratio was needed than in WT animals. This likely reflects a reduced fluidity in restricted membrane areas because of a higher enrichment in sphingolipids in the case of ASMKO brain.     

Chk2 phosphorylation of BRCA1 regulates DNA double-strand break repair

The pathway determining malignant cellular transformation, which depends upon mutation of the BRCA1 tumor suppressor gene, is poorly defined. A growing body of evidence suggests that promotion of DNA double-strand break repair by homologous recombination (HR) may be the means by which BRCA1 maintains genomic stability, while a role of BRCA1 in error-prone nonhomologous recombination (NHR) processes has just begun to be elucidated. The BRCA1 protein becomes phosphorylated in response to DNA damage, but the effects of phosphorylation on recombinational repair are unknown. In this study, we tested the hypothesis that the BRCA1-mediated regulation of recombination requires the Chk2- and ATM-dependent phosphorylation sites. We studied Rad51-dependent HR and random chromosomal integration of linearized plasmid DNA, a subtype of NHR, which we demonstrate to be dependent on the Mre11-Rad50-Nbs1 complex. Prevention of Chk2-mediated phosphorylation via mutation of the serine 988 residue of BRCA1 disrupted both the BRCA1-dependent promotion of HR and the suppression of NHR. Similar results were obtained when endogenous Chk2 kinase activity was inhibited by expression of a dominant-negative Chk2 mutant. Surprisingly, the opposing regulation of HR and NHR did not require the ATM phosphorylation sites on serines 1423 and 1524. Together, these data suggest a functional link between recombination control and breast cancer predisposition in carriers of Chk2 and BRCA1 germ line mutations. We propose a dual regulatory role for BRCA1 in maintaining genome integrity, whereby BRCA1 phosphorylation status controls the selectivity of repair events dictated by HR and error-prone NHR.     

Comparative assessment of the cell-surface antigens and gene expression profiles of the gingival tissue biomarkers in subjects with fixed functional and removable functional orthodontic appliances

BackgroundThis study aimed to examine the cellular components of the gingiva during orthodontic treatment with fixed and removable appliances. The cellular and molecular cues of pathologies of the gingival tissue associated with the use of different orthodontic appliances could be studied.Materials and methodsTissue samples of gingiva were received from healthy patients undergoing gingivectomy for aesthetic purpose and from patients with fixed and removable functional orthodontic appliances undergoing gingivectomy for gingival overgrowth. The collected samples were stored in a sterile container with phosphate-buffered saline and to carry out further processes it was transported to the laboratory.ResultsCells positive for ECAD and NCAD were found to be increased in fixed appliances where as CD90 and CD105 positive cells showed no significant difference in all the three groups. CD24 and CD146 positive cells were increased significantly in removable and fixed than normal whereas CD133 positive cells were decreased in removable and fixed than normal. CD44 positive cells showed no noticeable change in all three groups. The gene expression levels of KRT5, SOX2, NANOG, and CXCL5 were found to be significantly increased in removable and fixed appliance groups. However, KRT8, CXCL10, and TIMP1 were increased only in fixed appliance group but CXCL10 showed decreased expression in removable appliance group. KRT6A, MYC, and MMP9 were decreased in fixed appliance group whereas MYC and MMP9 were increased in removable appliance group. KRT6A, KRT8, and TIMP1 showed no significant difference in removable appliance group.ConclusionThis study demonstrated essential roles of various genes, showing their contribution in regulating cell proliferation and migration in both the removable and fixed functional appliances.     

A biomimetic approach towards synthesis of zinc oxide nanoparticles

Using natural processes as inspiration, the present study demonstrates a positive correlation between zinc metal tolerance ability of a soil fungus and its potential for the synthesis of zinc oxide (ZnO) nanoparticles. A total of 19 fungal cultures were isolated from the rhizospheric soils of plants naturally growing at a zinc mine area in India and identified on the genus, respectively the species level. Aspergillus aeneus isolate NJP12 has been shown to have a high zinc metal tolerance ability and a potential for extracellular synthesis of ZnO nanoparticles under ambient conditions. UV–visible spectroscopy, Fourier transform infrared spectroscopy, X-ray diffraction analysis, transmission electron microscopy, and energy dispersive spectroscopy studies further confirmed the crystallinity, morphology, and composition of synthesized ZnO nanoparticles. The results revealed the synthesis of spherical nanoparticles coated with protein molecules which served as stabilizing agents. Investigations on the role of fungal extracellular proteins in the synthesis of nanoparticles indicated that the process is nonenzymatic but involves amino acids present in the protein chains.     

A Novel Cancer Testis Antigen,A-Kinase Anchor Protein 4 (AKAP4) Is a Potential Biomarker for Breast Cancer

Background

Breast cancer is the second leading cause of cancer related deaths in women worldwide. Reports about the early diagnosis of breast cancer are suggestive of an improved clinical outcome and overall survival rate in cancer patients. Therefore, cancer screening biomarker for early detection and diagnosis is urgently required for timely treatment and better cancer management. In this context, we investigated an association of cancer testis antigen, A-Kinase anchor protein 4 (AKAP4) with breast carcinoma.

Methodology/Findings

We first compared the AKAP4 gene and protein expression in four breast cancer cells (MCF7, MDA-MB-231, SK-BR3 and BT474) and normal human mammary epithelial cells. In addition, 91 clinical specimens of breast cancer patients of various histotypes including ductal carcinoma in situ, infiltrating ductal carcinoma and infiltrating lobular carcinoma and 83 available matched adjacent non-cancerous tissues were examined for AKAP4 gene and protein expression by employing in situ RNA hybridization and immunohistochemistry respectively. Humoral response against AKAP4 was also investigated in breast cancer patients employing ELISA. Our in vitro studies in all breast cancer cells revealed AKAP4 gene and protein expression whereas, normal human mammary epithelial cells failed to show any expression. Using in situ RNA hybridization and immunohistochemistry, 85% (77/91) tissue specimens irrespective of histotypes, stages and grades of breast cancer clinical specimens revealed AKAP4 gene and protein expression. However, matched adjacent non-cancerous tissues failed to display any AKAP4 gene and protein expression. Furthermore, humoral response was observed in 79% (72/91) of total breast cancer patients. Interestingly, we observed that 94% (72/77) of breast cancer patients found positive for AKAP4 protein expression generated humoral response against AKAP4 protein.

Conclusions

Collectively, our data suggests that AKAP4 may be used as serum based diagnostic test for an early detection and diagnosis of breast cancer and may be a potential target for immunotherapeutic use.     

Frequency of alleles conferring resistance to a Bacillus thuringiensis toxin in a Philippine population of Scirpophaga incertulas (Lepidoptera: Pyralidae)

Using the F2 screen methodology, we estimated the frequency of alleles conferring resistance to the Cry1Ab toxin of Bacillus thuringiensis Berliner in a Philippine population of the stem borer Scirpophaga incertulas (Walker). Evaluation of >450 isofemale lines for survival of F2 larvae on cry1Ab plants did not detect the presence of an allele conferring a high level of resistance. The frequency of such an allele in the sampled population was conservatively estimated to be <3.6 x 10(-3) with 95% confidence and a detection probability of 94%. However, there was evidence of the presence of alleles conferring partial resistance to Cry1Ab. The frequency of alleles for partial resistance was estimated as 4.8 x 10(-3) with a 95% CI between 1.3 x 10(-3) and 1.04 x 10(-2) and a detection probability of 94%. Our results suggest that the frequency of alleles conferring resistance to Cry1Ab in the population of S. incertulas sampled is not too high to preclude successful implementation of the high dose/refuge resistance management strategy.     

Characterization and phylogenetic analysis of an entomopathogenic Bacillus cereus strain WGPSB-2 (MTCC 7182) isolated from white grub, Anomala dimidiata (Coleoptera: Scarabaeidae)

White grubs (Coleoptera: Scarabaeidae) are cosmopolitan and polyphagous insect pests of agricultural crops, forests and pastures around the world. The lack of an environmentally sound approach for white grub management has prompted the exploration and detection of a novel microbial biocontrol agent against these sub-terranean insect pests. In this study we describe the isolation, establishment of pathogenesis, biochemical characterization and phylogenetic analysis of an entomopathogenic Bacillus cereus strain WGPSB-2 (MTCC 7182), isolated from an atrophied pupa of Anomala dimidiata, collected from the N.W. Indian Himalayas. The sequencing and subsequent comparison of the 16S rDNA revealed that the strain has100% similarity with Bacillus cereus sequences. Phylogenetic analysis of the 16S rDNA sequence revealed that the isolate is closely related to Bacillus thuringiensis and Bacillus sphaericus. In vitro bioassays showed that the isolate was able to infect and cause 92 and 67% mortality in second instar larvae of Anomala dimidiata and Holotrichia seticollis, respectively. The infected larvae exhibited bacterial septicemia like symptoms and mortality occurred between the third and ninth weeks after inoculation. The culture has been granted the accession number MTCC 7182 by the Microbial Type Culture Collection and Gene Bank, Institute of Microbial Technology, Chandigarh, India.