Effect of the CRAC Peptide, VLNYYVW, on mPTP Opening in Rat Brain and Liver Mitochondria

Abstract

The translocator protein (TSPO; 18 kDa) is a high-affinity cholesterol-binding protein located in the outer membrane of mitochondria. A domain in the C-terminus of TSPO was characterized as the cholesterol recognition/interaction amino acid consensus (CRAC). The ability of the CRAC domain to bind to cholesterol led us to hypothesize that this peptide may participate in the regulation of mitochondrial membrane permeability. Herein, we report the effect of the synthetic CRAC peptide, VLNYYVW, on mitochondrial permeability transition pore (mPTP) opening. It was found that the CRAC peptide alone prevents the mPTP from opening, as well as the release of apoptotic factors (cytochrome c, AIF, and EndoG) in rat brain mitochondria (RBM). Co-incubation of CRAC, together with the TSPO drug ligand, PK 11195, resulted in the acceleration of mPTP opening and in the increase of apoptotic factor release. VLNYYVW did not induce swelling in rat liver mitochondria (RLM). 3,17,19-androsten-5-triol (19-Atriol; an inhibitor of the cholesterol-binding activity of the CRAC peptide) alone and in combination with the peptide was able to stimulate RLM swelling, which was Ca2+- and CsA-sensitive. Additionally, a combination of 19-Atriol with 100 nM PK 11195 or with 100 µM PK 11195 displayed the opposite effect: namely, the addition of 19-Atriol with 100 µM PK 11195 in a suspension of RLM suppressed the Ca2+-induced swelling of RLM by 40%, while the presence of 100 nM PK 11195 with 19-Atriol enhanced the swelling of RLM by 60%. Taken together, these data suggest the participation of the TSPO’s CRAC domain in the regulation of permeability transition.

The Effects of PK11195 and Protoporphyrin IX Can Modulate Chronic Alcohol Intoxication in Rat Liver Mitochondria under the Opening of the Mitochondrial Permeability Transition Pore

“…Moreover, the effect of ligands may also depend on their concentration. Thus, we previously showed that different concentrations of PK11195 can have opposite effects on the mPTP [35,74], which is apparently associated with different mechanisms of action. The ligands in the nanomolar concentration range studied in this work are likely to have a greater specificity for the TSPO, while higher concentrations can lead to other, TSPO-independent effects of both PPIX and PK11195.…”

Modulation of the Cholesterol-Dependent Activity of Macrophages IC-21 by CRAC Peptides with Substituted Motif-Forming Amino Acids

“…It was previously shown that peptide VLNYYVW, a fragment of a CRAC motif without a cationic amino acid, cholesterol-dependently modulates the activity of cultured macrophages IC-21 [29][30][31]. According to Azarashvili et al [48], the same peptide prevents the opening of non-selective mitochondrial pores (mPTP) and the release of cytochrome c, as well as other apoptotic factors in mitochondria isolated from rat brain.…”

Translocator Protein 18 kDa (TSPO): A Promising Molecular Target for Image-Guided Surgery of Solid Cancers

“… 48 , 49 Moreover, its structure was composed of an integral membrane protein built by the five transmembrane alpha-helices, namely two intramitochondrial loops, two extramitochondrial loops, one extramitochondrial C-terminal, one intramitochondrial N-terminal, and one cholesterol-binding domain. 50 Previous studies reported the occurrence of TSPO as a monomer, but current investigations suggested that it could generate oligomeric aggregates with itself and other proteins. There was an indication that it could transform into homo-oligomers to provide a binding site for cholesterol.…”