POGLUT2 and POGLUT3 O-glucosylate multiple EGF repeats in fibrillin-1, -2, and LTBP1 and promote secretion of fibrillin-1

• Published in: The Journal of biological chemistry 2021-09, Vol.297 (3), p.101055-101055, Article 101055
• Main Authors: Williamson, Daniel B., Sohn, Camron J., Ito, Atsuko, Haltiwanger, Robert S.
• Format: Article
• Language: English
• Subjects: Amino Acid Motifs; EGF repeats; extracellular matrix protein; Fibrillin-1 - chemistry; Fibrillin-1 - genetics; Fibrillin-1 - metabolism; Fibrillin-2 - chemistry; Fibrillin-2 - genetics; Fibrillin-2 - metabolism; glycoprotein secretion; Glycosylation; Humans; Latent TGF-beta Binding Proteins - chemistry; Latent TGF-beta Binding Proteins - genetics; Latent TGF-beta Binding Proteins - metabolism; Marfan Syndrome - enzymology; Marfan Syndrome - genetics; Marfan Syndrome - metabolism; mass spectrometry; O-glucose; protein folding; Protein Translocation Systems; Signal Transduction
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/j.jbc.2021.101055

Fibrillin-1 (FBN1) is the major component of extracellular matrix microfibrils, which are required for proper development of elastic tissues, including the heart and lungs. Through protein–protein interactions with latent transforming growth factor (TGF) β-binding protein 1 (LTBP1), microfibrils regulate TGF-β signaling. Mutations within the 47 epidermal growth factor-like (EGF) repeats of FBN1 cause autosomal dominant disorders including Marfan Syndrome, which is characterized by disrupted TGF-β signaling. We recently identified two novel protein O-glucosyltransferases, Protein O-glucosyltransferase 2 (POGLUT2) and 3 (POGLUT3), that modify a small fraction of EGF repeats on Notch. Here, using mass spectral analysis, we show that POGLUT2 and POGLUT3 also modify over half of the EGF repeats on FBN1, fibrillin-2 (FBN2), and LTBP1. While most sites are modified by both enzymes, some sites show a preference for either POGLUT2 or POGLUT3. POGLUT2 and POGLUT3 are homologs of POGLUT1, which stabilizes Notch proteins by addition of O-glucose to Notch EGF repeats. Like POGLUT1, POGLUT2 and 3 can discern a folded versus unfolded EGF repeat, suggesting POGLUT2 and 3 are involved in a protein folding pathway. In vitro secretion assays using the N-terminal portion of recombinant FBN1 revealed reduced FBN1 secretion in POGLUT2 knockout, POGLUT3 knockout, and POGLUT2 and 3 double-knockout HEK293T cells compared with wild type. These results illustrate that POGLUT2 and 3 function together to O-glucosylate protein substrates and that these modifications play a role in the secretion of substrate proteins. It will be interesting to see how disease variants in these proteins affect their O-glucosylation.