Leukemia virus long terminal repeat activates NFκB pathway by a TLR3-dependent mechanism

AbstractThe long terminal repeat (LTR) region of leukemia viruses plays a critical role in tissue tropism and pathogenic potential of the viruses. We have previously reported that U3-LTR from Moloney murine and feline leukemia viruses (Mo-MuLV and FeLV) upregulates specific cellular genes in trans in an integration-independent way. The U3-LTR region necessary for this action does not encode a protein but instead makes a specific RNA transcript. Because several cellular genes transactivated by the U3-LTR can also be activated by NFκB, and because the antiapoptotic and growth promoting activities of NFκB have been implicated in leukemogenesis, we investigated whether FeLV U3-LTR can activate NFκB signaling. Here, we demonstrate that FeLV U3-LTR indeed upregulates the NFκB signaling pathway via activation of Ras-Raf-IκB kinase (IKK) and degradation of IκB. LTR-mediated transcriptional activation of genes did not require new protein synthesis suggesting an active role of the LTR transcript in the process. Using Toll-like receptor (TLR) deficient HEK293 cells and PKR−/− mouse embryo fibroblasts, we further demonstrate that although dsRNA-activated protein kinase R (PKR) is not necessary, TLR3 is required for the activation of NFκB by the LTR. Our study thus demonstrates involvement of a TLR3-dependent but PKR-independent dsRNA-mediated signaling pathway for NFκB activation and thus provides a new mechanistic explanation of LTR-mediated cellular gene transactivation

Genetic and molecular regulation of gamma globin gene expression in patients with sickle cell disease

Thesis (Ph.D.)--Boston UniversityPLEASE NOTE: Boston University Libraries did not receive an Authorization To Manage form for this thesis or dissertation. It is therefore not openly accessible, though it may be available by request. If you are the author or principal advisor of this work and would like to request open access for it, please contact us at [email protected]. Thank you.The variability of clinical severity in sickle cell anemia patients has been attributed in part to fetal hemoglobin (HbF) expression. Hereditary persistence of fetal hemoglobin (HPFH) describes benign disorders that are characterized by increased y-globin chain expression often with a reduction or absence of B-globin chain expression. HPFH can be due to naturally occurring deletions at the 3' end of the B-globin locus. Variable size deletions that remove alpha(HBO) and B(HBB) globin genes result in HPFH and alphaB-thalassemia. We examined clinical and hematology data in 28 patients with sickle hemoglobin (HbS)/HPFH. We found HbS/HPFH patients did not have anemia, and had slightly reduced mean corpuscular volume (MCV). Their age, hemoglobin and MCV were found to be correlated with HbF levels. These individuals were asymptomatic when compared to homozygous HbSS patients even with unusually high levels of HbF. Three major quantitative trait loci (QTL) significantly associated with HbF levels in individuals from different populations have been identified and include polymorphisms in the Gy-globin gene (HBG2) promoter, BCL 11A, and the HBS1 L-MYB intergenic region. We investigated polymorphisms in these QTL in a unique cohort of 20 African American patients with sickle cell anemia expressing HbF levels equal to or greater than 11%. We also found significant associations of HbF in 2 of the 3 major loci, BCL11A (rs766432) (P=0.05), and HBS1L-MYB intergenic region (rs9399137) (P= 0.02). A 3 basepair (bp) (TAG) deletion in high linkage disequilibrium with rs9399137 in the HBS1 L-MYB intergenic region might also account for high HbF expression. Two QTL influence HbF levels in African Americans with sickle cell anemia but together account for 20% of HbF variance [1]. Therefore to further explore possible causes of high HbF, we sequenced a 14.1 kilobases (kb) DNA fragment between Ay-globin gene (HBG1) and HBD in 15 high HbF and 15 low HbF patients. The DNA fragment houses the 7.2kb Corfu deletion that is associated with elevated HbF levels in the homozygous state and also contains binding sites for BCL11A. Thirty-eight single nucleotide polymorph isms (SNPs) were present in both groups of patients. Four SNPs had significantly higher major allele frequencies in the high HbF group (P<0.05) suggesting that polymorphisms in this area might contribute to elevated HbF levels in African American sickle cell anemia patients.2031-01-0

Fetal hemoglobin in sickle cell anemia

Fetal hemoglobin (HbF) is the major genetic modulator of the hematologic and clinical features of sickle cell disease, an effect mediated by its exclusion from the sickle hemoglobin polymer. Fetal hemoglobin genes are genetically regulated, and the level of HbF and its distribution among sickle erythrocytes is highly variable. Some patients with sickle cell disease have exceptionally high levels of HbF that are associated with the Senegal and Saudi-Indian haplotype of the HBB-like gene cluster; some patients with different haplotypes can have similarly high HbF. In these patients, high HbF is associated with generally milder but not asymptomatic disease. Studying these persons might provide additional insights into HbF gene regulation. HbF appears to benefit some complications of disease more than others. This might be related to the premature destruction of erythrocytes that do not contain HbF, even though the total HbF concentration is high. Recent insights into HbF regulation have spurred new efforts to induce high HbF levels in sickle cell disease beyond those achievable with the current limited repertory of HbF inducers