Abstract
Here, a large fraction of plant mitochondrial actin was found to be resistant to protease and high-salt treatments, suggesting it was protected by mitochondrial membranes. A portion of this actin became sensitive to protease or high-salt treatment after removal of the mitochondrial outer membrane, indicating that some actin is located inside the mitochondrial outer membrane. The import of an actin-green fluorescent protein (GFP) fusion protein into the mitochondria in a transgenic plant, actin:GFP, was visualized in living cells and demonstrated by flow cytometry and immunoblot analyses. Polymerized actin was found in mitochondria of actin:GFP plants and in mung bean (Vigna radiata). Notably, actin associated with mitochondria purified from early-developing cotyledons during seed germination was sensitive to high-salt and protease treatments. With cotyledon ageing, mitochondrial actin became more resistant to both treatments. The progressive import of actin into cotyledon mitochondria appeared to occur in concert with the conversion of quiescent mitochondria into active forms during seed germination. The binding of actin to mitochondrial DNA (mtDNA) was demonstrated by liquid chromatography-tandem mass spectrometry analysis. Porin and ADP/ATP carrier proteins were also found in mtDNA-protein complexes. Treatment with an actin depolymerization reagent reduced the mitochondrial membrane potential and triggered the release of cytochrome C. The potential function of mitochondrial actin and a possible actin import pathway are discussed.
Highlights
Bacteria were once thought to be shapeless, disorganized biomolecular sacks but are known to possess a complex subcellular organization created by cytoskeletal components that are homologous to eukaryotic actin, tubulin, and intermediate filaments (Lowe and Amos, 1998; Jones et al, 2001; van den Ent et al, 2001; Ausmees et al, 2003; Errington, 2003; Carballido-Lopez, 2006; Graumann, 2007)
Since actin is the most abundant protein in most cells, to demonstrate unambiguously that actin exists in mitochondria, it is critical to remove loosely bound and peripheral actin associated with the mitochondrial surface systematically
Without using any protein cross-linkers, we demonstrated that mitochondrial actin was bound to well-bound mitochondrial DNA (mtDNA), which represents the mtDNA-nucleoprotein complex still organized to its organelle form following pulsed-field gel electrophoresis (PFGE) fractionation of lysed intact mitochondria embedded in an agarose plug
Summary
Bacteria were once thought to be shapeless, disorganized biomolecular sacks but are known to possess a complex subcellular organization created by cytoskeletal components that are homologous to eukaryotic actin, tubulin, and intermediate filaments (Lowe and Amos, 1998; Jones et al, 2001; van den Ent et al, 2001; Ausmees et al, 2003; Errington, 2003; Carballido-Lopez, 2006; Graumann, 2007). The mitochondrion is a structurally and functionally well-organized organelle. Reyes et al (2011) demonstrated that b-actin and myosin resided in human mitochondria by applying protease protection and iodixanol gradient analysis, respectively, and by import studies. We communicate our findings on the localization and characterization of actin in plant mitochondria as
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
