Collapse of late endosomal pH elicits a rapid Rab7 response via the V-ATPase and RILP

Area of Science:

• Cell Biology
• Molecular Biology
• Biochemistry

Background:

• Endosomal-lysosomal trafficking requires endosomal acidification by H+-V-ATPase for proper lysosomal pH.
• Disrupted pH homeostasis impairs lysosomal function and proteostasis, affecting protein synthesis and degradation.
• Lysosomotropic compounds can neutralize endosomal pH, impacting cellular processes.

Purpose of the Study:

• To investigate the effects of the dipeptide LLOMe on late endosomes (LEs) and their associated trafficking pathways.
• To elucidate the molecular mechanisms by which LLOMe affects LE pH and Rab7 activation.
• To identify novel pathways regulating endosomal pH and Rab7 function.

Main Methods:

• Treatment of mammalian cells with LLOMe and ammonium chloride (NH4Cl) to induce pH neutralization.
• Analysis of Rab7 activation, endosomal tubulation, and CI-M6PR trafficking.
• Investigating the assembly of V-ATPase V1G1 subunit and its interaction with Rab7 and RILP.

Main Results:

• LLOMe neutralizes LE pH without lysosomal membrane permeabilization, leading to Rab7 hyperactivation.
• pH neutralization disrupts endosomal tubulation and CI-M6PR recycling.
• Increased V-ATPase V1G1 subunit assembly on endosomal membranes stabilizes GTP-bound Rab7 via RILP.

Conclusions:

• A novel pathway is proposed where V-ATPase and RILP cooperate to modulate LE pH and Rab7 activation.
• This pathway contributes to pH control during physiological endosomal maturation and starvation.
• Pathological pH neutralization, via lysosomotropic compounds or disease states, may involve this mechanism.