Dynamic changes in the higher-level chromatin organization of specific sequences revealed by in situ hybridization to nuclear halos

A novel approach to study the higher level packaging of specific DNA sequences has been developed by coupling high-resolution fluorescence hybridization with biochemical fractionation to remove histones and distend DNA loops to form morphologically reproducible nuclear halos. Results demonstrate consistent differences in the organization of specific sequences, and further suggest a relationship to functional activity. Pulse-incorporated bromodeoxyuridine representing nascent replicating DNA localized with the base of the chromatin loops in discrete clustered patterns characteristic of intact cells, whereas at increasing chase times, the replicated DNA was consistently found further out on the extended region of the halo. Fluorescence hybridization to unique loci for four transcriptionally inactive sequences produced long strings of signal extending out onto the DNA halo or loop, whereas four transcriptionally active sequences remained tightly condensed as single spots within the residual nucleus. In contrast, in non-extracted cells, all sequences studied typically remained condensed as single spots of fluorescence signal. Interestingly, two transcriptionally active, tandemly repeated gene clusters exhibited strikingly different packaging by this assay. Analysis of specific genes in single cells during the cell cycle revealed changes in packaging between S-phase and non S-phase cells, and further suggested a dramatic difference in the structural associations in mitotic and interphase chromatin. These results are consistent with and suggestive of a loop domain organization of chromatin packaging involving both stable and transient structural associations, and provide precedent for an approach whereby different biochemical fractionation methods may be used to unravel various aspects of the complex higher-level organization of the genome

Processing of endogenous pre-mRNAs in association with SC-35 domains is gene specific

Analysis of six endogenous pre-mRNAs demonstrates that localization at the periphery or within splicing factor-rich (SC-35) domains is not restricted to a few unusually abundant pre-mRNAs, but is apparently a more common paradigm of many protein-coding genes. Different genes are preferentially transcribed and their RNAs processed in different compartments relative to SC-35 domains. These differences do not simply correlate with the complexity, nuclear abundance, or position within overall nuclear space. The distribution of spliceosome assembly factor SC-35 did not simply mirror the distribution of individual pre-mRNAs, but rather suggested that individual domains contain both specific pre-mRNA(s) as well as excess splicing factors. This is consistent with a multifunctional compartment, to which some gene loci and their RNAs have access and others do not. Despite similar molar abundance in muscle fiber nuclei, nascent transcript trees of highly complex dystrophin RNA are cotranscriptionally spliced outside of SC-35 domains, whereas posttranscriptional tracks of more mature myosin heavy chain transcripts overlap domains. Further analyses supported that endogenous pre-mRNAs exhibit distinct structural organization that may reflect not only the expression and complexity of the gene, but also constraints of its chromosomal context and kinetics of its RNA metabolism

Influenza surveillance in 15 countries in Africa, 2006-2010

BACKGROUND: In response to the potential threat of an influenza pandemic, several international institutions and governments, in partnership with African countries, invested in the development of epidemiologic and laboratory influenza surveillance capacity in Africa. METHODS: We used a standardized form to collect information on influenza surveillance system characteristics, the number and percent of influenza-positive patients with influenza-like illness (ILI) or severe acute respiratory infections (SARI) and virologic data. RESULTS: Between 2006 and 2010, the number of ILI and SARI sites in 15 African countries increased from 21 to 127 and from 2 to 98, respectively. Influenza was detected in 22% of ILI cases and 10% of SARI cases. Children 0-4 years accounted for 48% all ILI and SARI cases of which 20% and 10 respectively were positive for influenza. Influenza peaks were generally discernible in North and South Africa. Substantial co-circulation of influenza A and B occurred most years. CONCLUSIONS: Influenza is a major cause of respiratory illness in Africa, especially in children. Further strengthening influenza surveillance, along with conducting special studies on influenza burden, cost of illness, and role of other respiratory pathogens will help detect novel influenza viruses and inform and develop targeted influenza prevention policy decisions in the region.The work presented in this manuscript was funded completely or in part by host governments, Institute Pasteur, and cooperative agreements with the U.S. Centers for Disease Control and Prevention and/or the U.S. Department of Defense.http://www.journals.uchicago.edu/toc/jid/currenthb2013ay201

Nonrandom gene organization: structural arrangements of specific pre-mRNA transcription and splicing with SC-35 domains

Abstract. This work demonstrates a highly nonrandom distribution of specific genes relative to nuclear domains enriched in splicing factors and poly(A) ÷ RNA, and provides evidence for the direct involvement of these in pre-mRNA metabolism. As investigated in hundreds of diploid fibroblasts, human collagen Ietl and 13-actin DNA/RNA showed a very high degree of spatial association with SC-35 domains, whereas three nontranscribed genes, myosin heavy chain, neurotensin, and albumin, showed no such preferential association. Collagen Itxl RNA accumulates within the more central region of the domain, whereas [3-actin RNA localizes at the periphery. A novel approach revealed that collagen RNA tracks are polarized, with the entire gene at one end, on the edge of the domain, an

Repositioning of Muscle-specific Genes Relative to the Periphery of SC-35 Domains during Skeletal Myogenesis

Previous studies have shown that in a given cell type, certain active genes associate with SC-35 domains, nuclear regions rich in RNA metabolic factors and excluded from heterochromatin. This organization is not seen for all active genes; therefore, it is important to determine whether and when this locus-specific organization arises during development and differentiation of specific cell types. Here, we investigate whether gene organization relative to SC-35 domains is cell type specific by following several muscle and nonmuscle genes in human fibroblasts, committed but proliferative myoblasts, and terminally differentiated muscle. Although no change was seen for other loci, two muscle genes (Human β-cardiac myosin heavy chain and myogenin) became localized to the periphery of an SC-35 domain in terminally differentiated muscle nuclei, but not in proliferative myoblasts or in fibroblasts. There was no apparent change in gene localization relative to either the chromosome territory or the heterochromatic compartment; thus, the gene repositioning seemed to occur specifically with respect to SC-35 domains. This gene relocation adjacent to a prominent SC-35 domain was recapitulated in mouse 3T3 cells induced into myogenesis by introduction of MyoD. Results demonstrate a cell type-specific reorganization of specific developmentally regulated loci relative to large domains of RNA metabolic factors, which may facilitate developmental regulation of genome expression

Multiple‐victim

Previous studies of homicides in South Africa have examined serial murders and mass killings. While some scholars have examined parricides in African countries such as Ghana and Zimbabwe, few have examined the intersection of parricide and multiple victim homicides in the context of South Africa. This paper examines multiple victim parricides in a South African context using newspapers and court records as data. Eighteen cases of multiple‐victim parricides were identified and analysed. Our findings indicate that multiple victim parricides in South Africa are shaped by residential patterns as well as social and cultural factors that are unique to South Africa that are embodied in the offence characteristics. The implications on the findings are discussed.University of Pretoria's Visiting Scholarhttp://wileyonlinelibrary.com/journal/jip2021-08-03hj2020Early Childhood Educatio

Clonal Selection for Transcriptionally Active Viral Oncogenes during Progression to Cancer

Primary keratinocytes immortalized by human papillomaviruses (HPVs), along with HPV-induced cervical carcinoma cell lines, are excellent models for investigating neoplastic progression to cancer. By simultaneously visualizing viral DNA and nascent viral transcripts in interphase nuclei, we demonstrated for the first time a selection for a single dominant papillomavirus transcription center or domain (PVTD) independent of integrated viral DNA copy numbers or loci. The PVTD did not associate with several known subnuclear addresses but was almost always perinucleolar. Silent copies of the viral genome were activated by growth in the DNA methylation inhibitor 5-azacytidine. HPV-immortalized keratinocytes supertransduced with HPV oncogenes and selected for marker gene coexpression underwent crisis, and the surviving cells transcribed only the newly introduced genes. Thus, transcriptional selection in response to environmental changes is a dynamic process to achieve optimal gene expression for cell survival. This phenomenon may be critical in clonal selection during carcinogenesis. Examination of HPV-associated cancers supports this hypothesis

Influenza Surveillance in 15 Countries in Africa, 2006–2010

International audienceBACKGROUND: In response to the potential threat of an influenza pandemic, several international institutions and governments, in partnership with African countries, invested in the development of epidemiologic and laboratory influenza surveillance capacity in Africa and the African Network of Influenza Surveillance and Epidemiology (ANISE) was formed. METHODS: We used a standardized form to collect information on influenza surveillance system characteristics, the number and percent of influenza-positive patients with influenza-like illness (ILI), or severe acute respiratory infection (SARI) and virologic data from countries participating in ANISE. RESULTS: Between 2006 and 2010, the number of ILI and SARI sites in 15 African countries increased from 21 to 127 and from 2 to 98, respectively. Children 0-4 years accounted for 48% of all ILI and SARI cases of which 22% and 10%, respectively, were positive for influenza. Influenza peaks were generally discernible in North and South Africa. Substantial cocirculation of influenza A and B occurred most years. CONCLUSIONS: Influenza is a major cause of respiratory illness in Africa, especially in children. Further strengthening influenza surveillance, along with conducting special studies on influenza burden, cost of illness, and role of other respiratory pathogens will help detect novel influenza viruses and inform and develop targeted influenza prevention policy decisions in the region