Endocytic Trafficking and Human Diseases

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Mucolipins constitute a family of endosomal cation channels with homology to the transient receptor potential superfamily. In mammals, the mucolipin family includes three members, mucolipin-1, -2, and -3 (MCOLN1-3). MCOLN1 is the best-characterized member of the family due to the fact that mutations in this protein are associated with a human disease known as mucolipidosis type IV (MLIV). MLIV is an autosomal recessive disease characterized by mental and psychomotor retardation, diminished muscle tone or hypotonia, achlorhydria, and visual problems including corneal clouding, retinal degeneration, sensitivity to light, and strabismus. Analysis of fibroblasts from MLIV patients by electron microscopy revealed the presence of enlarged vacuolar structures that accumulate mucopolysaccharides and lipids forming characteristic multiconcentric lamellae. These enlarged vacuoles are present not only in fibroblasts but in every tissue and organ of MLIV patients, suggesting a general impairment of the lysosomal function. Our previous studies revealed that MCOLN1 mediates transient, regulated, and localized calcium efflux from late endosomes and lysosomes. Release of luminal calcium plays a pivotal role in several trafficking events, including fusion of lysosomes with autophagosomes, late endosomes, and plasma membrane. In agreement with this idea we found that MCOLN1 is required for proper membrane trafficking along the endosomal pathway and maintenance of ion homeostasis. Althoug the use of primary cells and cell lines has provided crucial information for understanding the function of MCOLN1, it will be of great importance to further validate these studies done in vitro with in vivo observations. For this reason, we are currently working in the generation of MCOLN1 knockouts in zebrafish. The use of zebrafish as a potentially useful model for our ongoing studies is based on a number of features, including the fact that they are easy to maintain in large numbers in a small laboratory space, their generation time is relatively short, and they present unique advantages for embryological analysis. Specifically we are working in the induction of acute MCOLN1 depletion by injection with specific morpholionos as well as in the generation of stable genetically engineered MCOLN1-knockout zebrafish by the use of Zinc Finger Nucleases.