VMA21 deficiency prevents vacuolar ATPase assembly and causes autophagic vacuolar myopathy.

X-linked Myopathy with Excessive Autophagy (XMEA) is a childhood onset disease characterized by progressive vacuolation and atrophy of skeletal muscle. We show that XMEA is caused by hypomorphic alleles of the VMA21 gene, that VMA21 is the diverged human ortholog of the yeast Vma21p protein, and that like Vma21p, VMA21 is an essential assembly chaperone of the vacuolar ATPase (V-ATPase), the principal mammalian proton pump complex. Decreased VMA21 raises lysosomal pH which reduces lysosomal degradative ability and blocks autophagy.

VMA21 deficiency causes an autophagic myopathy by compromising V-ATPase activity and lysosomal acidification.

X-linked myopathy with excessive autophagy (XMEA) is a childhood-onset disease characterized by progressive vacuolation and atrophy of skeletal muscle. We show that XMEA is caused by hypomorphic alleles of the VMA21 gene, that VMA21 is the diverged human ortholog of the yeast Vma21p protein, and that like Vma21p it is an essential assembly chaperone of the V-ATPase, the principal mammalian proton pump complex. Decreased VMA21 raises lysosomal pH, which reduces lysosomal degradative ability and blocks autophagy.

Differential intracellular transport and binding of verotoxin 1 and verotoxin 2 to globotriaosylceramide-containing lipid assemblies.

Although verotoxin-1 (VT1) and verotoxin-2 (VT2) share a common receptor, globotriaosyl ceramide (Gb(3)), VT2 induces distinct animal pathology and is preferentially associated with human disease. Moreover VT2 cytotoxicity in vitro is less than VT1. We therefore investigated whether these toxins similarly traffic within cells via similar Gb(3) assemblies.

Lipid metabolism and dynamics during phagocytosis.

Phagocytosis, the engulfment of particles, mediates the elimination of invading pathogens as well as the clearance of apoptotic cells. Ingested particles reside within a vacuole or phagosome, where they are eventually destroyed and digested. The phagosomal lumen acquires microbicidal and digestive properties through interaction with various components of the endocytic pathway, a process known as maturation.

Quantitative and dynamic assessment of the contribution of the ER to phagosome formation.

Phagosomes were traditionally thought to originate from an invagination and scission of the plasma membrane to form a distinct intracellular vacuole. An alternative model implicating the endoplasmic reticulum (ER) as a major component of nascent and maturing phagosomes was recently proposed (Gagnon et al., 2002).

The nature of the phagosomal membrane: endoplasmic reticulum versus plasmalemma.

For decades, the vacuole that surrounds particles engulfed by phagocytosis was believed to originate from the plasma membrane. Conversion of the nascent phagosome into a microbicidal organelle was thought to result from the subsequent, orderly fusion of early endosomes, late endosomes, and ultimately, lysosomes with the original plasma membrane-derived vacuole. This conventional model has been challenged, if not superseded, by a revolutionary model that regards phagosome formation as resulting from the particle sliding into the endoplasmic reticulum via an opening at the base of the phagocytic cup.

The pH of the secretory pathway: measurement, determinants, and regulation.

The luminal pH of the secretory pathway plays a critical role in the posttranslational modification and sorting of proteins and lipids. The pH of each one of the organelles that constitute the pathway is unique, becoming more acidic as the biosynthetic cargo approaches its destination. The methods used for measurement of pH in the secretory pathway, its determinants, and its regulation are the subjects of this review.

Disruption of a salt bridge dramatically accelerates subunit exchange in duck delta2 crystallin.

Intragenic complementation is a unique property of oligomeric enzymes with which to study subunit-subunit interactions. Complementation occurs when different subunits, each possessing distinct mutations that render the individual homomutant proteins inactive, interact to form a heteromutant protein with partial recovery of activity. In this paper, complementation events between human argininosuccinate lyase (ASL) and its homolog, duck delta2 crystallin, were characterized.

Molecular and cellular mechanisms underlying iron transport deficiency in microcytic anemia.

A mutation of the iron transporter Nramp2 (DMT1, Slc11a2) causes microcytic anemia in mk mice and in Belgrade rats by impairing iron absorption in the duodenum and in erythroid cells, causing severe iron deficiency. Both mk and Belgrade animals display a glycine-to-arginine substitution at position 185 (G185R) in the fourth predicted transmembrane domain of Nramp2. To study the molecular basis for the loss of function of Nramp2(G185R), we established cell lines stably expressing extracellularly tagged versions of wild-type (WT) or mutated transporters.