Cytoskeleton and regulation of mitochondrial function: the role of beta-tubulin II

Abstract

The control of mitochondrial function is a cardinal issue in the field of cardiac bioenergetics, and the analysis of mitochondrial regulations is central to basic research and in the diagnosis of many diseases. Interaction between cytoskeletal proteins and mitochondria can actively participate in mitochondrial regulation. Potential candidates for the key roles in this regulation are the cytoskeletal proteins plectin and tubulin. Analysis of cardiac cells has revealed regular arrangement of β-tubulin II, fully co-localized with mitochondria. β-Tubulin IV demonstrated a characteristic staining of branched network, β-tubulin III was matched with Z-lines, and β-tubulin I was diffusely spotted and fragmentary polymerized. In contrast, HL-1 cells were characterized by the complete absence of β-tubulin II. Comparative analysis of cardiomyocytes and HL-1 cells revealed a dramatic difference in the mechanisms of mitochondrial regulation. In the heart, colocalization of β-tubulin isotype II with mitochondria suggests that it can participate in the coupling of ATP-ADP translocase (ANT), mitochondrial creatine kinase (MtCK), and VDAC (ANT-MtCK-VDAC). This mitochondrial supercomplex is responsible for the efficient intracellular energy transfer via the phosphocreatine pathway. Existing data suggest that cytoskeletal proteins may control the VDAC, contributing to maintenance of mitochondrial and cellular physiology.

Keywords: HL-1 cells; VDAC; beta tubulin isotypes; cardiomyocytes; confocal microscopy; creatine kinase; mitochondria-cytoskeleton interactions; mitochondrial regulation.

Figure 1

(A) Complex and multiple role of the cytoskeleton in mitochondrial regulations under normal and pathological conditions. SR, sarcoplasmic reticulum. (B) Possible interactions of porin (VDAC) of the mitochondrial outer membrane (MOM). IFs, intermediate filaments; IMM, the inner mitochondrial membrane; IMS, inter-membrane space; Mito CK, mitochondrial creatine kinase. Tubulin controls VDAC permeability for ADP and ATP. LP is a still unknown cytoskeletal linker protein, which may also interact with VDAC and tubulin to regulate permeability of the MOM.

Figure 2

(A) Distribution of total β-tubulins (microtubular network), (B) β-Tubulin IV. (C) The mitochondria-specific isoform β-tubulin II stained with anti-β-tubulin II antibody followed by FITC secondary antibodies and demonstrating a typical mitochondria-like staining/arrangement in isolated adult rat heart cardiomyocytes (comparable to D). (D) Imaging of mitochondria in the same cells stained with fluorescent mitochondria-specific probe TMRM.