Abstract
ABSTRACTIn Drosophila, formation of the cardiac extracellular matrix (ECM) starts during embryogenesis. Assembly and incorporation of structural proteins such as Collagen IV, Pericardin, and Laminin A, B1, and B2 into the cardiac ECM is critical to the maintenance of heart integrity and functionality and, therefore, to longevity of the animal. The cardiac ECM connects the heart tube with the alary muscles; thus, the ECM contributes to a flexible positioning of the heart within the animal's body. Moreover, the cardiac ECM holds the larval pericardial nephrocytes in close proximity to the heart tube and the inflow tract, which is assumed to be critical to efficient haemolymph clearance. Mutations in either structural ECM constituents or ECM receptors cause breakdown of the ECM network upon ageing, with disconnection of the heart tube from alary muscles becoming apparent at larval stages. Finally, the heart becomes non-functional. Here, we characterised existing and new pericardin mutants and investigated biosynthesis, secretion, and assembly of Pericardin in matrices. We identified two new pericardin alleles, which turned out to be a null (pericardin3-548) and a hypomorphic allele (pericardin3-21). Both mutants could be rescued with a genomic duplication of a fosmid coding for the pericardin locus. Biochemical analysis revealed that Pericardin is highly glycosylated and forms redox-dependent multimers. Multimer formation is remarkably reduced in animals deficient for the prolyl-4 hydroxylase cluster at 75D3-4.
Highlights
By underlying or encasing a multitude of cells or tissues, extracellular matrices (ECMs) are essential to several physiological processes including tissue protection, tissue scaffolding, and cell signalling
We found that Pericardin processing, i.e. multimerisation, is not blocked in mutants for pdi and dplod, and – to some extent – is inhibited in deficiencies that delete a cluster of PH4-encoding genes, which is in contrast to Collagen IV processing phenotypes seen in mutants for pdi, dplod, or PH4 genes (Abrams and Andrew, 2002; Bunt et al, 2011; Kelemen-Valkony et al, 2012; Molnar et al, 2005; Myllyharju and Kivirikko, 2004; Pastor-Pareja and Xu, 2011; Yasothornsrikul et al, 1997)
Identification of two new pericardin mutants For screening a collection of pupal lethal EMS-induced mutants (Koundakjian et al, 2004), we made use of our previously established cardiac hand-GFP reporter to monitor age-related cardiac integrity defects in living animals without the need of dissection (Paululat and Heinisch, 2012; Sellin et al, 2006) (Fig. 1A). This led to the identification of two new mutant alleles carrying EMS-induced mutations on the third chromosome, PMM3-21 and PMM3-548, with both displaying an irreversible detachment of the pericardial nephrocytes from the heart tube eminent in third instar larvae (Fig. 1B,C)
Summary
By underlying or encasing a multitude of cells or tissues, extracellular matrices (ECMs) are essential to several physiological processes including tissue protection, tissue scaffolding, and cell signalling. University of Osnabrü ck, Biology, Department of Zoology and Developmental Biology, Barbarastraße 11, 49076 Osnabrü ck, Germany. The complexity of matrices is reflected by the number of proteins that constitute the matrix, and by the different ratio with which the various components contribute and by the appearance of unique components in matrices of specific tissues. Within the meshwork of its structural components, the ECM is spatially patterned and thereby provides locally restricted reaction environments and structural micro-compartments
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