Monkey Kidney Cells with mEmerald-α-Actinin and mCherry-PDHA1
alpha-actinin is an actin cross-linking protein with multiple roles in different cell types. In cardiac cells, isoforms are localized to the Z disc (the region of the sarcomere into which thin filaments are inserted) and analogous dense bodies, where they help anchor the myofibrillar actin filaments. alpha-actinin consists of an N-terminal actin binding domain, a central rod domain, and a C-terminal domain. The distance between cross-linked actin filaments is determined by the rod domain and also works as an interaction site for cytoskeletal and signaling proteins.
PDHA1 is a pyruvate dehydrogenase enzyme that is useful in microscopy applications as a mitochondrial marker. Along with dihydrolipoyl transacetylase (DLAT) and dihydrolipoyl dehydrogenase (DLD), PDHA1 forms a pyruvate dehydrogenase complex that functions in the catalysis of the irreversible conversion of pyruvate to acetyl-CoA. PDHA1 is heterotetramer comprised of two alpha and two beta subunits.
The dynamic behavior of mitochondria was visualized in this digital video with the red fluorescent protein mCherry fused to PDHA1. An original member of the mFruit series of fluorescent proteins, mCherry was produced via the directed evolution of mRFP1. Excitation and emission maxima of mCherry occur at 587 and 610 nanometers, respectively. The featured African green monkey kidney fibroblast cells (CV-1 line) are also shown expressing mEmerald fused to alpha-actinin, highlighting stress fibers and other actin-rich cellular structures.