Eliosin, A Protein Encoded by a Transcript from the HmPKD Locus, Is a Component of Mitochondria-ER Contact Sites/Mitochondria-Associated Membranes (MAMs)
- Genetic Diseases: Cystic - Basic
November 03, 2023 | Location: Exhibit Hall, Pennsylvania Convention Center
Abstract Time: 10:00 AM - 12:00 PM
Category: Genetic Diseases of the Kidneys
- 1201 Genetic Diseases of the Kidneys: Cystic
- Bacallao, Robert L., Richard L Roudebush VA Medical Center, Indianapolis, Indiana, United States
- Lazar, Virginie, Indiana University School of Medicine, Indianapolis, Indiana, United States
- Xu, Weimin, Indiana University School of Medicine, Indianapolis, Indiana, United States
The HmPKD1 gene locus is predicted to produce multiple transcripts whose functions are largely unknown. We studied one alternative transcript that starts at intron 40 but has a protein start site in exon 41. This transcript has a splice from the 3’ end of exon 41 to the 5’ end of exon 43 of the HmPKD1 gene. Subsequent splices follow the same splicing pattern found in the full-length polycystin-1 mRNA. The subcellular localization and function of this alternative transcript is not known.
PCR primers are designed to detect the transcript's unique splice features that are not found in full-length HmPKD1 transcripts. RT-PCR studies were conducted to identify the alternative transcript. cDNA encoding the alternative transcript and its product (Eliosin) were expressed in COS-1, 293, and NIH 3T3 cells for immune-blot and light microscopy analysis. Co-localization studies were performed with mCherry-Eliosin fusion protein and dynamin-related protein (DRP-1) or mitofusin-1 (MFN-1). Analysis of mitochondria morphology in co-transfection studies was performed in 293 and PKD 9-7 cells.
RT-PCR reactions performed using human kidney RNA confirmed that a shortened PCR fragment from the PKD1 gene’s exons 41/43 splice is expressed. Immune blot analysis revealed that the protein, we name Eliosin is a 48 kDa protein and it is expressed by a cDNA isolated from human testes. Fluorescence microscopy studies show co-localization between Eliosin and Inositol-3-phosphate receptor or MFN-1, known components of MAMs. However, when Eliosin and DRP-1 are co-expressed, DRP-1 is found in the cytosol. Since DRP-1 mediates mitochondria scission, we reasoned that the mutation in Eliosin leads to unopposed mitochondria scission in PKD 9-7 cells. We find that untransfected PKD 9-7 cells have fragmented mitochondria while Eliosin-transfected PKD 9-7 cells have normal-appearing mitochondria.
Eliosin is a 48 kDa protein that is a component of mitochondria-ER membrane contact sites, and it acts to displace dynamin-related protein-1 from MAMs. We conclude that Eliosin plays a role in altering the balance between mitochondria fusion and scission. This finding extends the HmPKD1 locus’ role in mitochondria metabolic physiology.
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