Effect of Antiretroviral Therapy on Apoptosis Markers and Morphology in Peripheral Lymph Nodes of HIV-Infected Individuals

Background:: CD4+ T cell depletion and destruction and the involution of the lymphoid tissue are hallmarks of HIV infection. Although the underlying mechanisms are still unclear, apoptosis appears to play a central role. The objective of this study was to investigate the effect of antiretroviral therapy on the lymph node tissue, particularly with respect to morphology and apoptosis. Patients and Methods:: Between 1997 and 1999, two inguinal lymph nodes were excised from 31 previously untreated individuals who were in an early stage of HIV infection, the first one prior to treatment and the second after 16 to 20 months of treatment. Paraffin sections were investigated for lymph node architecture, distribution of cellular and viral markers, apoptosis, and expression of apoptotic key molecules which indirectly reflect apoptotic processes. Results:: After 16-20 months of antiretroviral therapy, a significant decrease in highly activated HIV-driven immune response was observed in the lymph node tissue as a marked reduction in follicular hyperplasia, a normalization of the follicular dendritic cell network, a significant increase in the number of CD4+ T cells, and a significant decrease in the number of CD8+ T cells. The expression of several proapoptotic (Fas, TRAIL, and active caspase 3) and antiapoptotic (Bcl-2 and IL-7Rα) molecules that were reconstituted in the tissues during therapy resembled their expression in lymph nodes of HIV-negative individuals. Limitations of the study are (a) the lack of untreated patients in the late stages, (b) for ethical reasons, the lack of a control group with untreated patients, and (c) for methodological reasons, the restriction of sequential measurements of apotpotic markers to one-third of the patients. Conclusion:: Antiretroviral therapy initiated in the early stages in HIV infection may halt the irreversible destruction of the lymph node tissue and may partially normalize apoptotic processe

Evaluation of techniques for performing cellular isolation and preservation during microgravity conditions

Genomic and epigenomic studies require the precise transfer of microliter volumes among different types of tubes in order to purify DNA, RNA, or protein from biological samples and subsequently perform analyses of DNA methylation, RNA expression, and chromatin modifications on a genome-wide scale. Epigenomic and transcriptional analyses of human blood cells, for example, require separation of purified cell types to avoid confounding contributions of altered cellular proportions, and long-term preservation of these cells requires their isolation and transfer into appropriate freezing media. There are currently no protocols for these cellular isolation procedures on the International Space Station (ISS). Currently human blood samples are either frozen as mixed cell populations (within the CPT collection tubes) with poor yield of viable cells required for cell-type isolations, or returned under ambient conditions, which requires timing with Soyuz missions. Here we evaluate the feasibility of translating terrestrial cell purification techniques to the ISS. Our evaluations were performed in microgravity conditions during parabolic atmospheric flight. The pipetting of open liquids in microgravity was evaluated using analog-blood fluids and several types of pipette hardware. The best-performing pipettors were used to evaluate the pipetting steps required for peripheral blood mononuclear cell (PBMC) isolation following terrestrial density-gradient centrifugation. Evaluation of actual blood products was performed for both the overlay of diluted blood, and the transfer of isolated PBMCs. We also validated magnetic purification of cells. We found that positive-displacement pipettors avoided air bubbles, and the tips allowed the strong surface tension of water, glycerol, and blood to maintain a patent meniscus and withstand robust pipetting in microgravity. These procedures will greatly increase the breadth of research that can be performed on board the ISS, and allow improvised experimentation by astronauts on extraterrestrial missions

Biologia molecular aplicada à ciência das plantas daninhas.

The weeds have high genetic variability, mainly for this, they are adapted to environments disturbed by humans. Although weed control has evolved positively in recent years, they continue to interfere with agricultural production. The objective of this bibliographic review is to present the contribution of molecular biology in the studies applied to herbology. There area gap in what we learn about weed genomics, which could help us manage and improve the competitiveness of weed crops. Many studies in the field of weed science can be carried out using molecular biology techniques. The characterization of the genome of weed species, aiming to identify species with higher accuracy, identification of herbicide resistant species and its mechanism of resistance, genetic variability and similarity among weed populations, identification of genes involved in the interaction processes between plants, among other.As plantas daninhas possuem elevada variabilidade genética, e principalmente por este motivo, são adaptadas a ambientes com intensa atividade humana. Embora o controle de plantas daninhas tenha evoluído de maneira positiva nos últimos anos, elas continuam a interferir na produção agrícola. O objetivo desta revisão bibliográfica é apresentar a contribuição da biologia molecular nos estudos aplicados a herbologia. Há lacunas entre o que aprendemos sobre genômica de plantas daninhas e como esses conhecimentos poderiam nos auxiliar no manejo e melhorar a competividade de culturas agrícolas frente às plantas daninhas. Muitos estudos na área da ciência das plantas daninhas podem ser realizados com o emprego de técnicas de biologia molecular, sendo eles: caracterização do genoma de espécies de plantas daninhas, visando à identificação destes com maior acurácia, identificação de espécies resistentes a herbicidas e seu mecanismo de resistência, variabilidade e similaridade genética entre populações de plantas daninhas, identificação de genes envolvidos nos processos de interação entre plantas, dentre outros

Genetic dissimilarity in conyza sumatrensis revealed by simple sequence repeat (SSR) markers.

In view of the rapid evolution of Conyza sumatrensis populations resistant to glyphosate, it is necessary to understand the genetic diversity aimed to improve strategies for managing this weed. We investigated the genetic dissimilarity among 15 biotypes of C. sumatrensis from different geographic regions using microsatellite loci. The biotypes, were cultivated in a greenhouse to obtain vegetal material for DNA extraction. Nineteen microsatellite markers (SSR), were developed for C. sumatrensis biotypes. The genetic dissimilarity was estimated by the Jaccard coefficient (JC) and the biotypes grouped by the UPGMA method. The results demonstrated a high dissimilarity (JC = 7.14 to 82.62) of the analyzed material, with the biotypes forming five groups, being one group formed just by the susceptible biotype and in the others grouped by biotypes from distinct locations in the same group The high genetic diversity of C. sumatrensis indicates that the biotypes may show different responses to different management strategies, and that the mechanisms of resistance to herbicides and characteristics of evolution of populations due to adaptability may be some of the factors involved in the genetic variability of the species. Keywords: polymorphism; SSR; tall fleabane; genetic variability RESUMO: Tendo em vista a rápida evolução das populações de Conyza sumatrensis resistentes ao glifosato, é necessário entender a diversidade genética com vistas a melhorar as estratégias de manejo dessa planta daninha. Diante do exposto, objetivou-se com este trabalho investigar a dissimilaridade genética entre 15 biótipos de C. sumatrensis de diferentes regiões geográficas usando marcadores moleculares microssatélites. Os biótipos foram cultivados em casa de vegetação, para obtenção de material vegetal para extração de DNA. Dezenove marcadores microssatélites (SSR) foram desenvolvidos para os biótipos de C. sumatrensis. A dissimilaridade genética foi estimada pelo coeficiente de Jaccard (JC), e os biótipos, agrupados pelo método UPGMA. Os resultados demonstraram alta dissimilaridade (JC = 7,14 a 82,62) do material analisado, com os biótipos formando cinco grupos. A alta diversidade genética de C. sumatrensis indica que os biótipos podem apresentar distintas respostas a diferentes estratégias de manejo e que os mecanismos de resistência a herbicidas podem ser um dos fatores envolvidos na variabilidade genética das espécies. Palavras-chave: polimorfismo; SSR; buva; variabilidade genétic

CDK1-dependent Inhibition of the E3 Ubiquitin Ligase CRL4 CDT2 Ensures Robust Transition from S Phase to Mitosis

Replication-coupled destruction of a cohort of cell cycle proteins ensures efficient and precise genome duplication. Three proteins destroyed during replication via the CRL4CDT2 ubiquitin E3 ligase, CDT1, p21, and SET8 (PR-SET7), are also essential or important during mitosis, making their reaccumulation after S phase a critical cell cycle event. During early and mid-S phase and during DNA repair, proliferating cell nuclear antigen (PCNA) loading onto DNA (PCNADNA) triggers the interaction between CRL4CDT2 and its substrates, resulting in their degradation. We have discovered that, beginning in late S phase, PCNADNA is no longer sufficient to trigger CRL4CDT2-mediated degradation. A CDK1-dependent mechanism that blocks CRL4CDT2 activity by interfering with CDT2 recruitment to chromatin actively protects CRL4CDT2 substrates. We postulate that deliberate override of replication-coupled destruction allows anticipatory accumulation in late S phase. We further show that (as for CDT1) de novo SET8 reaccumulation is important for normal mitotic progression. In this manner, CDK1-dependent CRL4CDT2 inactivation contributes to efficient transition from S phase to mitosis

Drosophila Symplekin localizes dynamically to the histone locus body and tricellular junctions

The scaffolding protein Symplekin is part of multiple complexes involved in generating and modifying the 3' end of mRNAs, including cleavage-polyadenylation, histone pre-mRNA processing and cytoplasmic polyadenylation. To study these functions in vivo, we examined the localization of Symplekin during development and generated mutations of the Drosophila Symplekin gene. Mutations in Symplekin that reduce Symplekin protein levels alter the efficiency of both poly A+ and histone mRNA 3' end formation resulting in lethality or sterility. Histone mRNA synthesis takes place at the histone locus body (HLB) and requires a complex composed of Symplekin and several polyadenylation factors that associates with the U7 snRNP. Symplekin is present in the HLB in the early embryo when Cyclin E/Cdk2 is active and histone genes are expressed and is absent from the HLB in cells that have exited the cell cycle. During oogenesis, Symplekin is preferentially localized to HLBs during S-phase in endoreduplicating follicle cells when histone mRNA is synthesized. After the completion of endoreplication, Symplekin accumulates in the cytoplasm, in addition to the nucleoplasm, and localizes to tricellular junctions of the follicle cell epithelium. This localization depends on the RNA binding protein ypsilon schachtel. CPSF-73 and a number of mRNAs are localized at this same site, suggesting that Symplekin participates in cytoplasmic polyadenylation at tricellular junctions