Mitochondrial Alterations Induced by the p13II Protein of Human T-cell Leukemia Virus Type 1

Abstract

Human T-cell leukemia virus type 1 encodes a number of "accessory" proteins of unclear function; one of these proteins, p13 II , is targeted to mitochondria and disrupts mitochondrial morphology. The present study was undertaken to unravel the function of p13 II through (i) determination of its submitochondrial localization and sequences required to alter mitochondrial morphology and (ii) an assessment of the biophysical and biological properties of synthetic peptides spanning residues 9 -41 (p13 9 -41 ), which include the amphipathic mitochondrialtargeting sequence of the protein. p13 9 -41 folded into an ␣ helix in micellar environments. Fractionation and immunogold labeling indicated that full-length p13 II accumulates in the inner mitochondrial membrane. p13 9 -41 induced energy-dependent swelling of isolated mitochondria by increasing inner membrane permeability to small cations (Na ؉ , K ؉ ) and released Ca 2؉ from Ca 2؉ -preloaded mitochondria. These effects as well as the ability of full-length p13 II to alter mitochondrial morphology in cells required the presence of four arginines, forming the charged face of the targeting signal. The mitochondrial effects of p13 9 -41 were insensitive to cyclosporin A, suggesting that full-length p13 II might alter mitochondrial permeability through a permeability transition pore-independent mechanism, thus distinguishing it from the mitochondrial proteins Vpr and X of human immunodeficiency virus type 1 and hepatitis B virus, respectively.HTLV-1 1 is a complex retrovirus that is associated with two distinct pathologies, a leukemia/lymphoma of mature CD4 ϩ

The human T-cell leukemia virus type 1 p13II protein: effects on mitochondrial function and cell growth

“…including the MTS) revealed that it folds into an a-helix upon exposure to membrane-mimetic solutions containing phospholipids or sodium dodecyl sulfate (SDS) micelles. 31 As expected, a peptide in which the four arginines were substituted with four prolines, which are known to disrupt a-helical folding, failed to adopt an a-helical conformation, while substitution of the arginines with glutamines or alanines and leucines did not interfere with folding. 31 Surprisingly, none of these substitutions had a substantial effect on the ability of p13 II to accumulate in mitochondria (see below).…”

“…31 As expected, a peptide in which the four arginines were substituted with four prolines, which are known to disrupt a-helical folding, failed to adopt an a-helical conformation, while substitution of the arginines with glutamines or alanines and leucines did not interfere with folding. 31 Surprisingly, none of these substitutions had a substantial effect on the ability of p13 II to accumulate in mitochondria (see below). 31 Mitochondria are complex organelles bounded by two highly specialized membranes that define four mitochondrial subcompartments -outer membrane, intermembrane space, inner membrane, and matrix.…”

“…22 In addition, the observations made for the arginine substitution mutants described above indicate that the p13 II MTS does not require the presence of these positively charged residues and works independently of its ability to fold into an a-helix. 31 These properties do not HTLV-1 p13 II and mitochondria DM D'Agostino et al appear to fit well with any of the inner membrane protein import pathways described above and suggest that the p13 II MTS has peculiar sequence-structure requirements that might direct mitochondrial import through an alternative mechanism.…”

“…31 Surprisingly, none of these substitutions had a substantial effect on the ability of p13 II to accumulate in mitochondria (see below). 31 Mitochondria are complex organelles bounded by two highly specialized membranes that define four mitochondrial subcompartments -outer membrane, intermembrane space, inner membrane, and matrix. Immunoelectron microscopy and fractionation techniques based on differential sensitivity to extraction with digitonin and sodium carbonate demonstrated that p13 II is an integral membrane protein and accumulates mainly in the inner mitochondrial membrane.…”

“…Immunoelectron microscopy and fractionation techniques based on differential sensitivity to extraction with digitonin and sodium carbonate demonstrated that p13 II is an integral membrane protein and accumulates mainly in the inner mitochondrial membrane. 31 Import of nuclear-encoded proteins into mitochondria involves interactions with specific chaperones and translocases in the outer and inner membranes, and proceeds through different pathways depending on the nature of the protein's targeting sequence and structure (reviewed in Wiedemann et al 32 ). Some inner membrane proteins contain an N-terminal positively charged, cleavable presequence and a hydrophobic sequence that stops the protein in the inner membrane.…”