Reduction of Lysyl Hydroxylase 3 Causes Deleterious Changes in the Deposition and Organization of Extracellular Matrix

Maija Risteli,Heli Ruotsalainen,Antti M Salo,Raija Sormunen,Laura Sipilä,Naomi L Baker,Shireen R Lamandé,Leena Vimpari-Kauppinen,Raili Myllylä

doi:10.1074/jbc.m109.038190

Abstract

Lysyl hydroxylase 3 (LH3) is a multifunctional enzyme possessing lysyl hydroxylase, collagen galactosyltransferase, and glucosyltransferase (GGT) activities. We report here an important role for LH3 in the organization of the extracellular matrix (ECM) and cytoskeleton. Deposition of ECM was affected in heterozygous LH3 knock-out mouse embryonic fibroblasts (MEF(+/-)) and in skin fibroblasts collected from a member of a Finnish epidermolysis bullosa simplex (EBS) family known to be deficient in GGT activity. We show the GGT deficiency to be due to a transcriptional defect in one LH3 allele. The ECM abnormalities also lead to defects in the arrangement of the cytoskeleton in both cell lines. Ultrastructural abnormalities were observed in the skin of heterozygous LH3 knock-out mice indicating that even a moderate decrease in LH3 has deleterious consequences in vivo. The LH3 null allele in the EBS family member and the resulting abnormalities in the organization of the extracellular matrix, similar to those found in MEF(+/-), may explain the correlation between the severity of the phenotype and the decrease in GGT activity reported in this family.

Highlights

28204 JOURNAL OF BIOLOGICAL CHEMISTRY mutations in the LH2 gene (PLOD2) that result in the complete absence of telopeptide hydroxylysine residues in bone collagens [16, 17]
We have examined heterozygous Lysyl hydroxylase 3 (LH3) knock-out mice and mouse embryonic fibroblasts (MEFϩ/Ϫ) at the ultrastructural level as well as cells from a human model of LH3 deficiency, an epidermolysis bullosa simplex (EBS) family member known to exhibit a decrease in galactosylhydroxylysyl glucosyltransferase (GGT) activity
When the gold particles in a 4.4-␮m2 area of the endoplasmic reticulum were counted in 30 cells, fewer particles, about 57% of controls, were observed in the EBS fibroblasts, revealing a reduction in the amount of LH3 in these cells. These results indicate that there are no molecular weight differences between the LH3 proteins produced by the EBS patient and control cells, the quantity of LH3, measured either as GGT activity or as LH3 protein, was decreased in the EBS patient and the decrease was more pronounced in the cell culture medium

Summary

Introduction

28204 JOURNAL OF BIOLOGICAL CHEMISTRY mutations in the LH2 gene (PLOD2) that result in the complete absence of telopeptide hydroxylysine residues in bone collagens [16, 17]. We have examined heterozygous LH3 knock-out mice and mouse embryonic fibroblasts (MEFϩ/Ϫ) at the ultrastructural level as well as cells from a human model of LH3 deficiency, an epidermolysis bullosa simplex (EBS) family member known to exhibit a decrease in GGT activity.

Results

Conclusion