Human long-chain acyl-coa synthetase 5 (acsl5) variantsfrom gene to pathology
- Ndagire, Dorothy
- Antonio Alcina Madueño Director
- Fuencisla Matesanz del Barrio Co-director
Defence university: Universidad de Granada
Fecha de defensa: 24 May 2012
- José Antonio López Guerrero Chair
- Ana Clara Abadía Molina Secretary
- José Mario Sabio Sánchez Committee member
- Alberto Fraile Ramos Committee member
- Francisco Martínez Abarca Committee member
- María Cristina Hernández López Committee member
Type: Thesis
Abstract
Background: Alternative promoter usage and polymorphisms in ACSL5 gene promoter may represent candidates for altering gene expression and thus causing complex diseases. The relationship of lipid metabolism and immune pathologies remains unclear although there are evidences that show that an interrupted process of lipid metabolism is implicated in multiple sclerosis (MS) and systemic lupus erythematosus (SLE).We investigated the association of ACSL with MS and SLE. Results: The search of ACSL5 transcript variants using the 5'-RACE (Rapid amplification of 5' cDNA ends) identified 6 alternative promoters from which ACSL5 is transcribed, labelled as 1A-1A', 1A', 1B, 1C, 1D and 2 in Jurkat T cells. Promoter 1A and 1A' are functional in K562 cells and not in lymphocytes. The transcript variants differed from each other only in the 5' sequences and no differences were observed from exon 3 to exon 21. Fluctuations of transcription start sites (TSS) were observed at the initial of exons 1B, 1C and 2. Luciferase reporter analysis and electrophoresis mobility shift assay (EMSA) of promoter 1C, the most active in lymphocytes, revealed a region of 100bp with maximal promoter activity which possessed functional DNA binding sequence for SP1 transcription factor. To determine the involvement of ACSL in SLE, we measured transcripts of ACSL isoforms in peripheral blood mononuclear cells (PBMCs) in patients with SLE and health controls using quantitative real-time-PCR (qRT-PCR) which revealed higher ACSL5 transcript levels blood cells of SLE patients than healthy controls. Furthermore, in vitro experiments showed that increased ACSL5 mRNA expression is correlated with increase in apoptosis and expression of apoptotic associated genes FAS, FASLG and TNF cytokine gene. Genotyping of polymorphism rs2419621 and rs2419629 located in ACSL5 gene in healthy controls and MS patients showed that the ACSL5 gene is associated with the disease. Conclusion: The ACSL5 gene transcription shows great isoform diversity leading to the different TSS that have specific cell-type expression preference. We localized the major proximal promoter controlling the expression of ACSL5 in activated Jurkat T cells and showed specific binding of SP1 transcription factor to this DNA region. ACSL5 is associated with MS, SLE and has a role in the apoptosis of lymphocytes.