Role of let-7b in the regulation of N-acetylgalactosaminyltransferase 2 in IgA nephropathy.
BACKGROUND
IgA nephropathy (IgAN) is characterized by aberrant O-glycosylation in the hinge region of IgA1. The early step in O-glycan formation is the attachment of N-acetylgalactosamine (GalNAc) to the serine/threonine of the hinge region; the process is catalysed by UDP-N-acetyl-α-d-galactosamine:polypeptide N-acetylgalactosaminyltransferase 2 (GALNT2). In our previous work, the microarray analysis on peripheral blood mononuclear cells (PBMCs) identified an upregulated miRNA called let-7b.


METHODS
To study the molecular mechanisms in which let-7b was involved, we performed a bioinformatic analysis to predict their target genes. To validate biologically let-7b targets, we performed transient transfection experiments ex vivo using PBMCs from an independent group of IgAN patients and healthy blood donors (HBDs).


RESULTS
Bioinformatic analysis revealed that GALNT2 is the potential target of let-7b. We found this miRNA significantly upregulated in PBMCs of IgAN patients compared with HBDs. Then, we demonstrated in ex-vivo experiments that let-7b decreased GALNT2 levels in PBMCs of IgAN patients, whereas the loss of let-7b function in PBMCs of HBDs led to an increase of GALNT2 mRNA and its protein level. Finally, we found that upregulation of let-7b occurred also in B-lymphocytes from IgAN patients.


CONCLUSIONS
Our results give novel additional information on the abnormal O-glycosylation process of IgA1 in IgAN patients. This study provides evidence for another important miRNA-based regulatory mechanism of the O-glycosylation process in which the deregulated expression of let-7b is associated with altered expression of GALNT2. This finding could be taken into consideration for new therapeutic approaches in IgAN because other serum glycosylated proteins do not display abnormal glycosylation.
Department of Emergency and Organ Transplantation, University of Bari, Bari, Italy C.A.R.S.O. Consortium, University of Bari, Valenzano (Ba), Italy.
Grazia
G
Serino
C.A.R.S.O. Consortium, University of Bari, Valenzano (Ba), Italy Dipartimento di Scienze e Tecnologie Biologiche ed Ambientali (DiSTeBA), Università del Salento, Monteroni, Lecce, Italy.
Fabio
F
Sallustio
C.A.R.S.O. Consortium, University of Bari, Valenzano (Ba), Italy.
Claudia
C
Curci
Department of Emergency and Organ Transplantation, University of Bari, Bari, Italy.
Sharon N
SN
Cox
Department of Emergency and Organ Transplantation, University of Bari, Bari, Italy Cardiovascular Genetics and Genomics, National Heart and Lung Institute, Imperial College London, London, UK.
Francesco
F
Pesce
C.A.R.S.O. Consortium, University of Bari, Valenzano (Ba), Italy.
Giuseppe
G
De Palma
C.A.R.S.O. Consortium, University of Bari, Valenzano (Ba), Italy Schena Foundation, Research Center for Kidney Diseases, Valenzano (Ba), Italy.
Francesco P
FP
Schena
D000328:Adult; D001402:B-Lymphocytes / Q000378:metabolism; D015415:Biomarkers / Q000378:metabolism; D015153:Blotting, Western; D016022:Case-Control Studies; D005260:Female; D005922:Glomerulonephritis, IGA / Q000201:enzymology* / Q000235:genetics* / Q000473:pathology; D006031:Glycosylation; D006801:Humans; D007668:Kidney / Q000378:metabolism*; D007963:Leukocytes, Mononuclear / Q000378:metabolism; D008297:Male; D035683:MicroRNAs / Q000235:genetics*; D017350:N-Acetylgalactosaminyltransferases / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014407:Tumor Cells, Cultured; D000097763:Polypeptide N-acetylgalactosaminyltransferase
D015415:Biomarkers; D035683:MicroRNAs; D017350:N-Acetylgalactosaminyltransferases; D012333:RNA, Messenger; D000097763:Polypeptide N-acetylgalactosaminyltransferase; C487931:mirnlet7 microRNA, human
Iga nephropathy; O-glycosylation process; UDP-N-acetyl-α-d-galactosamine:polypeptide N-acetylgalactosaminyltransferase 2 (GALNT2); microRNAs; peripheral blood mononuclear cells