A RSC/nucleosome complex determines chromatin architecture and facilitates activator binding.

How is chromatin architecture established and what role does it play in transcription? We show that the yeast regulatory locus UASg bears, in addition to binding sites for the activator Gal4, sites bound by the RSC complex. RSC positions a nucleosome, evidently partially unwound, in a structure that facilitates Gal4 binding to its sites. The complex comprises a barrier that imposes characteristic features of chromatin architecture. In the absence of RSC, ordinary nucleosomes encroach over the UASg and compete with Gal4 for binding. Taken with our previous work, the results show that both prior to and following induction, specific DNA-binding proteins are the predominant determinants of chromatin architecture at the GAL1/10 genes. RSC/nucleosome complexes are also found scattered around the yeast genome. Higher eukaryotic RSC lacks the specific DNA-binding determinants found on yeast RSC, and evidently Gal4 works in those organisms despite whatever obstacle broadly positioned nucleosomes present.

Endonuclease mediated genome editing in drug discovery and development: promises and challenges.

Site specific genome editing has been gradually employed in drug discovery and development process over the past few decades. Recent development of CRISPR technology has significantly accelerated the incorporation of genome editing in the bench side to bedside process. In this review, we summarize examples of applications of genome editing in the drug discovery and development process. We also discuss current hurdles and solutions of genome editing.

Predictors of Adolescent Internalizing and Externalizing Mental Health Symptoms: A School-Based Study in Southern India.

INTRODUCTION: Mental illnesses are one of the major contributors to the overall burden of disease among the young. We investigated the predictors of emotional and behavioral problems among in-school adolescents in the Indian context. METHODS: Using stratified sampling, 1441 adolescents were recruited to participate in the study in Udupi taluk. The study instruments included a socio-demographic pro forma and the adolescent self-reporting version of the Strengths and Difficulties Questionnaire (SDQ) to assess the emotional and behavioral problems among them. We explored the predictors of total difficulties, as well as externalizing and internalizing problems and gender differences. SPSS version 25 was used to analyze the data. Descriptive statistics, a Chi-square test for associations, an independent t-test to explore the gender differences, correlation analysis, and backward stepwise logistic regression for the predictors were used. RESULTS: The mean age of the participants was 15.31 ± 0.76. An almost equal percentage of male (49.6%) and female (50.4%) participants provided data. Abnormal scores were highest under conduct problems (8.5%), and the total difficulties reached 5.1%. The male participants had higher levels of conduct, hyperactivity, peer relationship, and externalizing problems the while the female participants experienced higher levels of emotional and internalizing problems. It was observed that there was a significant positive relationship between age and emotional problems, conduct problems, hyperactivity, peer problems, and total SDQ score. An older age predicted an abnormal total difficulties score and externalizing and internalizing behaviors, while the type of school predicted the total difficulties and internalizing behaviors. CONCLUSION: The age of the adolescent, their gender, and the type of school they attended emerged as predictors of the emotional and behavioral problems among them.

Activator control of nucleosome occupancy in activation and repression of transcription.

The relationship between chromatin structure and gene expression is a subject of intense study. The universal transcriptional activator Gal4 removes promoter nucleosomes as it triggers transcription, but how it does so has remained obscure. The reverse process, repression of transcription, has often been correlated with the presence of nucleosomes. But it is not known whether nucleosomes are required for that effect. A new quantitative assay describes, for any given location, the fraction of DNA molecules in the population that bears a nucleosome at any given instant. This allows us to follow the time courses of nucleosome removal and reformation, in wild-type and mutant cells, upon activation (by galactose) and repression (by glucose) of the GAL genes of yeast. We show that upon being freed of its inhibitor Gal80 by the action of galactose, Gal4 quickly recruits SWI/SNF to the genes, and that nucleosome "remodeler" rapidly removes promoter nucleosomes. In the absence of SWI/SNF, Gal4's action also results in nucleosome removal and the activation of transcription, but both processes are significantly delayed. Addition of glucose to cells growing in galactose represses transcription. But if galactose remains present, Gal4 continues to work, recruiting SWI/SNF and maintaining the promoter nucleosome-free despite it being repressed. This requirement for galactose is obviated in a mutant in which Gal4 works constitutively. These results show how an activator's recruiting function can control chromatin structure both during gene activation and repression. Thus, both under activating and repressing conditions, the activator can recruit an enzymatic machine that removes promoter nucleosomes. Our results show that whereas promoter nucleosome removal invariably accompanies activation, reformation of nucleosomes is not required for repression. The finding that there are two routes to nucleosome removal and activation of transcription-one that requires the action of SWI/SNF recruited by the activator, and a slower one that does not-clarifies our understanding of the early events of gene activation, and in particular corrects earlier reports that SWI/SNF plays no role in GAL gene induction. Our finding that chromatin structure is irrelevant for repression as studied here-that is, repression sets in as efficiently whether or not promoter nucleosomes are allowed to reform-contradicts the widely held, but little tested, idea that nucleosomes are required for repression. These findings were made possible by our nucleosome occupancy assay. The assay, we believe, will prove useful in studying other outstanding issues in the field.

p53 blocks RuvAB promoted branch migration and modulates resolution of Holliday junctions by RuvC.

The Holliday junction is the central intermediate in homologous recombination. Branch migration of this four-stranded DNA structure is a key step in genetic recombination that affects the extent of genetic information exchanged between two parental DNA molecules. Here, we have constructed synthetic Holliday junctions to test the effects of p53 on both spontaneous and RuvAB promoted branch migration as well as the effect on resolution of the junction by RuvC. We demonstrate that p53 blocks branch migration, and that cleavage of the Holliday junction by RuvC is modulated by p53. These findings suggest that p53 can block branch migration promoted by proteins such as RuvAB and modulate the cleavage by Holliday junction resolution proteins such as RuvC. These results suggest that p53 could have similar effects on eukaryotic homologues of RuvABC and thus have a direct role in recombinational DNA repair.