Membrane environment exerts an important influence on rac-mediated activation of phospholipase Cγ2

We performed analyses of the molecular mechanisms involved in the regulation of phospholipase Cγ2 (PLCγ2). We identified several regions in the PLCγ-specific array, γSA, that contribute to autoinhibition in the basal state by occlusion of the catalytic domain. While the activation of PLCγ2 by Rac2 requires stable translocation to the membrane, the removal of the domains required for membrane translocation in the context of an enzyme with impaired autoinhibition generated constitutive, highly active PLC in cells. We further tested the possibility that the interaction of PLCγ2 with its activator protein Rac2 was sufficient for activation through the release of autoinhibition. However, we found that Rac2 binding in the absence of lipid surfaces was not able to activate PLCγ2. Together with other observations, these data suggest that an important consequence of Rac2 binding and translocation to the membrane is that membrane proximity, on its own or together with Rac2, has a role in the release of autoinhibition, resulting in interfacial activation.     

Targeted disruption of Ing2 results in defective spermatogenesis and development of soft-tissue sarcomas

ING2 (inhibitor of growth family, member 2) is a member of the plant homeodomain (PHD)-containing ING family of putative tumor suppressors. As part of mSin3A-HDAC corepressor complexes, ING2 binds to tri-methylated lysine 4 of histone H3 (H3K4me3) to regulate chromatin modification and gene expression. ING2 also functionally interacts with the tumor suppressor protein p53 to regulate cellular senescence, apoptosis and DNA damage response in vitro, and is thus expected to modulate carcinogenesis and aging. Here we investigate the developmental and physiological functions of Ing2 through targeted germline disruption. Consistent with its abundant expression in mouse and human testes, male mice deficient for Ing2 showed abnormal spermatogenesis and were infertile. Numbers of mature sperm and sperm motility were significantly reduced in Ing2(-/-) mice (～2% of wild type, P<0.0001 and ～10% of wild type, P<0.0001, respectively). Their testes showed degeneration of seminiferous tubules, meiotic arrest before pachytene stage with incomplete meiotic recombination, induction of p53, and enhanced apoptosis. This phenotype was only partially abrogated by concomitant loss of p53 in the germline. The arrested spermatocytes in Ing2(-/-) testes were characterized by lack of specific HDAC1 accumulation and deregulated chromatin acetylation. The role of Ing2 in germ cell maturation may extend to human ING2 as well. Using publicly available gene expression datasets, low expression of ING2 was found in teratozoospermic sperm (>3-fold reduction) and in testes from patients with defective spermatogenesis (>7-fold reduction in Sertoli-cell only Syndrome). This study establishes ING2 as a novel regulator of spermatogenesis functioning through both p53- and chromatin-mediated mechanisms, suggests that an HDAC1/ING2/H3K4me3-regulated, stage-specific coordination of chromatin modifications is essential to normal spermatogenesis, and provides an animal model to study idiopathic and iatrogenic infertility in men. In addition, a bona fide tumor suppressive role of Ing2 is demonstrated by increased incidence of soft-tissue sarcomas in Ing2(-/-) mice.     

MAGI: Methylation analysis using genome information

By incorporating annotation information into the analysis of next-generation sequencing DNA methylation data, we provide an improvement in performance over current testing procedures. Methylation analysis using genome information (MAGI) is applicable for both unreplicated and replicated data, and provides an effective analysis for studies with low sequencing depth. When compared with current tests, the annotation-informed tests provide an increase in statistical power and offer a significance-based interpretation of differential methylation.     

Buffalograss germplasm resistance to Blissus occiduus (Hemiptera: Lygaeidae)

Plant germplasm collections may offer genetic variability useful in identifying insect resistance. The goal of this project was to evaluate buffalograss genotypes [Buchlo? dactyloides (Nutt.) Engelm.] for resistance to the chinch bug, Blissus occiduus Barber (Hemiptera: Lygaeidae), and to relate resistance to ploidy level, chinch bug number, and pubescence. Forty-eight buffalograss genotypes from diverse geographic locations were evaluated in replicated studies under greenhouse conditions. Of the genotypes studied, four were highly resistant, 22 were moderately resistant, 19 were moderately susceptible, and three were highly susceptible to chinch bug damage. The mean number of chinch bugs was significantly different among the 48 genotypes. There was no significant correlation between chinch bug resistance and ploidy level or chinch bug resistance and pubescence. These results indicate the genetic source of resistance to chinch bugs exists in buffalograss germplasm. Highly resistant genotypes can be used in breeding programs to further improve buffalograss cultivars.     

Chinch bug-resistant buffalograss: an investigation of tolerance, antixenosis, and antibiosis

Choice and no-choice studies were conducted to determine the categories (antibiosis, antixenosis, and tolerance) of resistance of four buffalograsses (NE91-118, 'Bonnie Brae', 'Cody', and 'Tatanka') previously identified as resistant to the western chinch bug, Blissus occiduus Barber. Antibiosis studies found no significant differences in western chinch bug fecundity, nymphal development, or survival among the resistant and susceptible buffalograsses. Tolerance studies indicated that NE91-118, Cody, and Tatanka exhibited moderate-to-high levels of tolerance based on western chinch bug damage ratings and plant height, whereas Bonnie Brae exhibited moderate-to-low levels of tolerance. Choice studies indicated the presence of antixenosis in NE91-118, whereas Cody and Tatanka showed little or no antixenosis. Scanning electron microscopy was used to disclose morphological differences between NE91-118 (resistant) and '378' (susceptible). The epicuticular wax structures and trichome densities were similar between 378 and NE91-118, suggesting that morphological structures do not contribute to NE91-118 antixenosis.