Ehler-Danlos syndrome causing aortic carboxypeptidase-like protein mutations and cellular stress
Embargo Date
2027-10-27
OA Version
Citation
Abstract
The extracellular matrix (ECM) is composed of a network of proteins that create a structural scaffold for cells and regulate cellular behaviors. When ECM proteins are mutated, pathologies of the connective tissue may arise, such as Ehler-Danlos syndrome (EDS). Aortic carboxypeptidase-like protein (ACLP) is an ECM protein implicated in ECM remodeling and collagen fibrillogenesis. The mutation of the gene for ACLP, AEBP1, results in a specific variant of EDS called, “Classical-like type 2 EDS.” Prior work has shown that specific ACLP mutants can be retained in the endoplasmic reticulum (ER) due to the insertion or deletion of codons that alter the structure of the protein precursors, hindering their folding and processing in the ER. Protein accumulation in the ER can lead to ER stress and activation of the unfolded protein response (UPR), as well as the integrated stress response (ISR). Two particular ACLP mutants, Ins40 and Leu642Pro, that are retained in the ER are implicated in classical-like type 2 EDS. The goal of our research was to determine whether the ACLP mutations involved in the pathogenesis of clEDS2 resulted in the activation of cellular stress pathways and if the stress response pathways could be attenuated with a therapeutic intervention. Using bacterial transformation and Western blot, we confirmed the intracellular retention of ACLP mutants Ins40 and Leu642Pro and the secretion of ACLP mutants Cys581*, Arg631*, Asn236Ser, and Trp356*. Using immunofluorescence, we sought to identify if the ISR signaling cascade was activated when Ins40 and Leu642Pro were intracellularly retained. We administered an ISR-inhibitor, ISRIB, to the retained mutants and used Western blot to investigate whether their secretion pathways could be salvaged. These studies confirmed the work of prior bodies of research and further the exploration into understanding the mechanisms of ACLP secretion, cellular stress pathways, and how they are impacted by therapeutic intervention such as ISRIB.
Description
2024