Exon 4-encoded sequence is a major determinant of cytotoxicity of apolipoprotein L1.
The apolipoprotein L1 (APOL1) gene (APOL1) product is toxic to kidney cells, and its G1 and G2 alleles are strongly associated with increased risk for kidney disease progression in African Americans. Variable penetrance of the G1 and G2 risk alleles highlights the significance of additional factors that trigger or modify the progression of disease. In this regard, the effect of alternative splicing in the absence or presence of G1 or G2 alleles is unknown. In this study we investigated whether alternative splicing of non-G1, non-G2 APOL1 (APOL1 G0) affects its biological activity. Among seven APOL1 exons, exons 2 and 4 are differentially expressed in major transcripts. We found that, in contrast to APOL1 splice variants B3 or C, variants A and B1 demonstrate strong toxicity in human embryonic kidney (HEK293T) cells. Subsequently, we established that exon 4 is a major determinant of toxicity of variants A and B1 and that extracellular release of these variants is dispensable for their cytotoxicity. Although only variants A and B1 induced nuclear translocation of transcription factor EB (TFEB), a master regulator of lysosomal biogenesis and autophagy, exon 4-positive and -negative APOL1 variants stimulated perinuclear accumulation of unprocessed autophagosomes. Knockdown of endogenous TFEB did not attenuate APOL1 cytotoxicity, indicating that nuclear translocation of TFEB is dispensable for APOL1 toxicity. Our findings that a human podocyte cell line expresses exon 4-positive and -negative APOL1 transcripts suggest that these variants may play a differential role in podocyte pathology. In summary, we have identified exon 4 as a major determinant of APOL1 G0 cytotoxicity.
Meharry Medical College, Center for AIDS Health Disparities Research, Nashville, Tennessee;
Atanu K
AK
Khatua
Meharry Medical College, Center for AIDS Health Disparities Research, Nashville, Tennessee;
Amber M
AM
Cheatham
Rappaport Faculty of Medicine and Research Institute, Technion-Israel Institute of Technology, Rambam Medical Center, Haifa, Israel; and.
Etty D
ED
Kruzel
Feinstein Institute for Medical Research, Manhasset, New York.
Pravin C
PC
Singhal
Rappaport Faculty of Medicine and Research Institute, Technion-Israel Institute of Technology, Rambam Medical Center, Haifa, Israel; and.
Karl
K
Skorecki
Meharry Medical College, Center for AIDS Health Disparities Research, Nashville, Tennessee; wpopik@mmc.edu.
Waldemar
W
Popik
D021581:Active Transport, Cell Nucleus; D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000075944:Apolipoprotein L1; D001053:Apolipoproteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D001343:Autophagy; D001483:Base Sequence; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors / Q000235:genetics / Q000378:metabolism; D005091:Exons; D005786:Gene Expression Regulation; D057809:HEK293 Cells; D006801:Humans; D008075:Lipoproteins, HDL / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D008969:Molecular Sequence Data; D050199:Podocytes / Q000378:metabolism* / Q000473:pathology; D034622:RNA Interference; D012333:RNA, Messenger / Q000378:metabolism; D015398:Signal Transduction; D013329:Structure-Activity Relationship; D014158:Transcription, Genetic; D014162:Transfection
C108666:APOL1 protein, human; D000075944:Apolipoprotein L1; D001053:Apolipoproteins; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D008075:Lipoproteins, HDL; D012333:RNA, Messenger; C064972:TFEB protein, human
RNA splicing; apolipoprotein L1; autophagy; cytotoxicity; endocytosis