Sequence diversity of tubulin isotypes in regulation of the mitochondrial voltage-dependent anion channel

Abstract

The microtubule protein tubulin is a heterodimer comprising / subunits, in which each subunit features multiple isotypes in vertebrates. For example, seven -tubulin and eight -tubulin isotypes in the human tubulin gene family vary mostly in the length and primary sequence of the disordered anionic C-terminal tails (CTTs). The biological reason for such sequence diversity remains a topic of vigorous enquiry. Here, we demonstrate that it may be a key feature of tubulin's role in regulation of the permeability of the mitochondrial outer membrane voltagedependent anion channel (VDAC). Using recombinant yeast /-tubulin constructs with -CTTs, -CTTs, or both from various human tubulin isotypes, we probed their interactions with VDAC reconstituted into planar lipid bilayers. A comparative study of the blockage kinetics revealed that either -CTTs or -CTTs block VDAC pore and that the efficiency of blockage by individual CTTs spans two orders of magnitude, depending on the CTT isotype.-Tubulin constructs, notably 3, blocked VDAC most effectively. We quantitatively describe these experimental results using a physical model that accounts only for the number and distribution of charges in the CTT, and not for the interactions between specific residues on the CTT and VDAC pore. Based on these results, we speculate that the effectiveness of VDAC regulation by tubulin depends on the predominant tubulin isotype in a cell. Consequently, the fluxes of ATP/ADP http://www.jbc.org/cgi

Mitochondrial bioenergetic dysfunction in the D2.mdx model of Duchenne muscular dystrophy is associated with microtubule disorganization in skeletal muscle

“…While Ca 2+ stress has been proposed as a primary cause of swollen mitochondria and impaired oxidative phosphorylation in mdx mice [ 5 ], separate observations of altered microtubule networks [ 37 , 38 , 42 ] were also linked to disrupted cytosolic NADPH oxidase-induced ROS and Ca 2+ signaling [ 42 ]. An intriguing possibility of a mitochondrial link to microtubule disorganization emerges when considering the separate discoveries that tubulin components of microtubules can directly bind to VDAC on the outer mitochondrial membrane and decrease its permeability to ADP/ATP cycling [ 10 , 14 , 21 , 29 ]. Considering that pharmacological alterations of microtubule networks changes tubulin-VDAC interactions and ADP-dependent bioenergetics [ 13 ], it seems plausible that the distinct observations of disorganized microtubules and mitochondrial dysfunctions in D2.…”

“…Nevertheless, this observation does not rule out the possibility of other tubulin-VDAC interaction combinations. For example, while the CTT tail of α-tubulin has been shown to block the pore of VDAC, there is a greater affinity of the CTT tail on certain β isotypes, particularly βIII and VDAC1 [14]. In addition, it has been suggested that free βII tubulin binds VDAC in muscle-independent of heterodimeric tubulin-such that this free pool may be a distinct regulator of ADP/ATP cycling through VDAC [15].…”

“…The second objective was to explore whether this potential relationship was related to altered tubulin-VDAC binding stemming from disorganized microtubules. Specific attention was given to α-tubulin considering it binds various isotypes of β-tubulin as an α/β heterodimer with the CTT tail of both components having affinity for VDAC [ 10 , 14 – 16 ]. VDAC2 was selected given 1) its deficiency results in embryonic death thereby demonstrating its importance [ 17 ], 2) it has been proposed that VDAC2 may uniquely regulate the more efficient creatine-dependent mitochondrial phosphate shuttling mechanism [ 18 – 21 ], and 3) we have previously shown that tubulin-VDAC2 interactions are changed when microtubule organization is altered by paclitaxel [ 13 ].…”

The Utility of Nanopore Technology for Protein and Peptide Sensing

“…Rostovsteva and colleagues used a co‐localization assay to identify a likely sensor for the tubulin, which has disordered c‐terminal tails (CTT) that differ between isotypes. Each CTT produces a unique ion current signature based on the distribution of negative charges along the sequence . Several systems have been devised that anchor recognition sites within the pore, such that a portion of the anchor is inside the pore either through a specific biological recognition element (i.e., biotin–streptavidin), or by integrating binding sites to the pore that capture the natural ligand of the protein analyte .…”

Deletion of VDAC1 Hinders Recovery of Mitochondrial and Renal Functions After Acute Kidney Injury

“…Apoptosis is associated with increased expression of VDAC1 in the outer mitochondrial membrane and oligomerization of VDAC1 monomers to form large channels that allow for release of mitochondrial pro-apoptotic proteins into the cytoplasm where they mediate additional apoptotic events [10,[15][16][17]. Regulation of cell survival signals by VDAC1 occurs through its interactions with proteins of the Bcl-2 family, hexokinase, protein kinases, cytoskeletal proteins (tubulin, α-synuclein, plectin, and desmin), and mitochondrial lipids (cardiolipin and phoshatidylethanolamine) [16][17][18][19][20][21][22][23][24].…”