Study of LvsB in Dictyostelium discoideum provides insights into the Chediak-Higashi syndrome.

摘要

The Chediak-Higashi Syndrome is a disorder affecting lysosome biogenesis. At the cellular level, the Chediak-Higashi syndrome is characterized by the presence of grossly enlarged lysosomes in every tissue. Impaired lysosomal function in CHS patients results in many physiological problems, including immunodeficiency, albinism and neurological problems. The Chediak-Higashi syndrome is caused by the loss of a BEACH protein of unknown function named Lyst.;In this work, I have studied the function of the Dictyostelium LvsB protein, the ortholog of mammalian Lyst and a protein that is also important for lysosomal function. Using a knock-in approach we tagged LvsB with GFP and expressed it from its single chromosomal locus. GFP-LvsB was observed on endocytic and phagocytic compartments. Specific analysis of the endocytic compartments labeled by LvsB showed that they represented late lysosomes and postlysosomes. The analysis of LvsB-null cells revealed that loss of LvsB resulted in enlarged postlysosomes, in the abnormal localization of proton pumps on postlysosomes and their abnormal acidification. This work demonstrated that the abnormal postlysosomes in LvsB-null cells were produced by the inappropriate fusion of lysosomes with postlysosomal compartments.;Furthermore, this work provided the first evidence that LvsB is a functional antagonist of the GTPase Rab14 in vesicle fusion events. In particular, we demonstrated that reduction of Rab14 activity suppressed the LvsB-null phenotype by reducing the enlarged post-lysosomes and the enhanced rate of heterotypic fusion. In contrast, expression of an active form of Rab14 enhanced the LvsB-null phenotype by causing an even more severe enlargement of endosome size.;The results provided by this work support the model that LvsB and Lyst proteins act as negative regulators of fusion by limiting the heterotypic fusion of early with late compartments and antagonize Rab GTPases in membrane fusion. The LvsB localization studies and the functional assessment of the LvsB-null phenotype helped make unique contributions to the understanding of the molecular function of Lyst proteins.