Institution Granting Degree
University of Virginia
Nucleoside diphosphate kinase, NM23, Primary cilia, Microtubules, Lipids, NDPK
Cytosolic nucleoside diphosphate kinases (NDPKs) have been implicated in a variety of signaling pathways that occur at membranes, including those that control cell migration and spreading. This is particularly intriguing, as cytosolic NDPKs (NDPK A and NDPK B) are soluble proteins and do not have membrane-binding motifs, leading to the question: how do NDPK's participate in such a wide array of membrane signaling processes?
Our lab has shown that one portion of cytosolic NDPK is translocated to the ruffling plasma membrane upon activation of both receptor tyrosine kinases (RTKs) and G protein-coupled receptors (GPCRs) and that the Rac1 signaling pathway is responsible for that migration. Although NDPK does not bind directly to Rac1, it moves to the cell periphery in conjunction with Rac1. While investigating the association of cytosolic NDPK with the plasma membrane, we found that another pool of NDPK is bound to membrane vesicles that are associated with microtubules (Mt/Ves). A detailed study of this NDPK population shows that, unlike the pool that is involved in Rac1 signaling, NDPK's presence in these vesicles is not dependent on extracellular stimulation; rather, it is controlled by the nucleotide triphosphate to nucleotide diphosphate ratio ([NTP]/[NDP]), as evidenced by the effect of nucleotides on Mt/Ves isolated from fibroblasts. More importantly, purified and cytosolic NDPKs bind to both immobilized lipids and liposomes in a nucleotide-sensitive manner. This indicates that NDPK can bind directly to intracellular membrane compartments, most likely to provide CTP for phospholipid biosynthesis and GTP for the many small GTPases involved in microtubule-dependent traffic.
We also found that NDPK localizes to yet another microtubule-based cell compartment: the sensory primary cilium, an organelle implicated in many signaling pathways. NDPK enters the cilium during its development, when it reaches about 5.5 microns (or 24% of final primary cilia length) in A6 cells. In primary cilia NDPK is present in the soluble portion, or matrix, and in association with the membrane fraction. The function of NDPK within primary cilia is most likely to regenerate GTP for microtubule turnover and for signaling systems, making it an important contributor to primary cilia structure and function.