The BEACH Protein LvsB Is Localized on Lysosomes and Postlysosomes and Limits Their Fusion with Early Endosomes

Abstract

The Chediak-Higashi syndrome (CHS) is a genetic disorder caused by the loss of the BEACH protein Lyst. Impaired lysosomal function in CHS patients results in many physiological problems, including immunodeficiency, albinism and neurological problems. Dictyostelium LvsB is the ortholog of mammalian Lyst and is also important for lysosomal function. A knock-in approach was used to tag LvsB with green fluorescent protein (GFP) and express it from its single chromosomal locus. GFP-LvsB was observed on late lysosomes and postlysosomes. Loss of LvsB resulted in enlarged postlysosomes, in the abnormal localization of proton pumps on postlysosomes and their abnormal acidification. The abnormal postlysosomes in LvsB-null cells were produced by the inappropriate fusion of early endosomal compartments with postlysosomal compartments. The intermixing of compartments resulted in a delayed transit of fluid-phase marker through the endolysosomal system. These results support the model that LvsB and Lyst proteins act as negative regulators of fusion by limiting the heterotypic fusion of early endosomes with postlysosomal compartments.

Comparison of Dictyostelium LvsB and endosomal fission defect mutants support a fusion regulatory role for LvsB

“…Initial characterization of beige mice suggested that Lyst functions to limit the rate of homotypic lysosome fusion (Oliver and Essner, 1975). This model for Lyst function was subsequently supported by many studies in humans (Stinchcombe et al, 2000), Dictyostelium Kypri et al, 2007), mice (Willingham et al, 1981;Hammel et al, 1987;Hammel et al, 2010), cats (Collier et al, 1985) and Drosophila (Rahman et al, 2012). An alternative model suggested that Lyst functions to control lysosomal fission instead of fusion (Burkhardt et al, 1993).…”

“…Disruption of LvsB results in the accumulation of large acidic compartments, much like those observed in cells of CHS patients Wang et al, 2002). Characterization of the LvsB-null phenotype has provided definitive evidence of an important role for LvsB during progression of the endo-lysosomal pathway (Kypri et al, 2007). However, similar to the conundrum of deciphering Lyst function in other systems, studies in Dictyostelium have proposed both the fusion and fission models for LvsB function.…”

“…However, similar to the conundrum of deciphering Lyst function in other systems, studies in Dictyostelium have proposed both the fusion and fission models for LvsB function. Studies published by Harris and Kypri (Kypri et al, 2007) proposed that LvsB has a regulatory role in vesicle fusion. The fusion model for LvsB function was corroborated by recent evidence of a functional relationship between LvsB and the fusion regulatory GTPase Rab14 (Kypri et al, 2013).…”

“…Under normal conditions, lysosomes undergo homotypic fusion to exchange their contents, but are prevented from undergoing heterotypic fusion with post-lysosomes to preserve the identity of these distinct stages (Maniak, 2003). Owing to the coordination of lysosome to post-lysosome maturation, vacuolin is rarely observed on vesicular membranes until late stages of endosome maturation in wild-type cells, and the majority of vacuolin-labeled vesicles have a neutral luminal pH (Jenne et al, 1998;Kypri et al, 2007). In contrast, vacuolin frequently labels acidic vesicles that contain early endocytic markers in LvsB-null cells (Kypri et al, 2007).…”

“…Owing to the coordination of lysosome to post-lysosome maturation, vacuolin is rarely observed on vesicular membranes until late stages of endosome maturation in wild-type cells, and the majority of vacuolin-labeled vesicles have a neutral luminal pH (Jenne et al, 1998;Kypri et al, 2007). In contrast, vacuolin frequently labels acidic vesicles that contain early endocytic markers in LvsB-null cells (Kypri et al, 2007). Both of these observations are used to support a model where LvsB inhibits heterotypic fusion between lysosomes and post-lysosomes, and the absence of LvsB allows fusion of neutral vacuolin-labeled post-lysosomes with earlier acidic lysosomes to form hybrid compartments.…”