Constitutive Plasma Membrane Turnover in T-REx293 cells via Ordered Membrane Domain Endocytosis under Mitochondrial Control.

Clathrin/dynamin-independent endocytosis of ordered plasma membrane domains (rdered embrane omain ndocytosis, OMDE) can become massive in response to cytoplasmic Ca elevations, G protein activation by non-hydrolyzable GTP analogs, and enhanced oxidative metabolism. In patch-clamped murine bone marrow macrophages (BMMs), cytoplasmic succinate and pyruvate, but not β-hydroxybutyrate, induce OMDE of 75% of the plasma membrane within 2 min. The responses require palmitoylation of membrane proteins, being decreased by 70% in BMMs lacking the acyltransferase, DHHC5, by treatment with carnitine to shift long-chain acyl groups from cytoplasmic to mitochondrial acyl-CoAs, by bromopalmitate/albumin complexes to block DHHCs, and by the mitochondria-specific cyclosporin, NIM811, to block permeability transition pores that may release mitochondrial coenzyme A into the cytoplasm.

Longitudinal diffusion barriers imposed by myofilaments and mitochondria in murine cardiac myocytes.

Using optical and electrical methods, we document that diffusion in the cytoplasm of BL6 murine cardiomyocytes becomes restricted >20-fold as molecular weight increases from 30 to 2,000, roughly as expected for pores with porin channel dimensions. Bodipy-FL ATP diffuses >40-fold slower than in free water at 25°C. From several fluorophores analyzed, bound fluorophore fractions range from 0.

Pore-like diffusion barriers in murine cardiac myocytes.

Using both optical and electrical methods, we document that solute diffusion in the cytoplasm of BL6 murine cardiac myocytes becomes restricted >30-fold as molecular weight increases from 30 to 2000, roughly as expected for pores with dimensions of cardiac porin channels. The Bodipy-FL ATP analogue diffuses ∼50-fold slower in BL6 cardiac cytoplasm than in free water. From several fluorophores analyzed, our estimates of bound fluorophore fractions range from 0.

TMEM16F and dynamins control expansive plasma membrane reservoirs.

Cells can expand their plasma membrane laterally by unfolding membrane undulations and by exocytosis. Here, we describe a third mechanism involving invaginations held shut by the membrane adapter, dynamin. Compartments open when Ca activates the lipid scramblase, TMEM16F, anionic phospholipids escape from the cytoplasmic monolayer in exchange for neutral lipids, and dynamins relax.