Citron kinase
Citron Rho-interacting kinase is an enzyme that in humans is encoded by the CIT gene.
Structure
Citron is a 183 kDa protein that contains a C6H2 zinc finger, a PH domain, and a long coiled-coil forming region including 4 leucine zippers and a rho / rac binding site. It was discovered as a rho/rac effector in 1995, interacting only with the GTP bound forms of rho and rac 1. Displaying a distinctive protein organization, this protein defines a separate class of rho partners. Using a cloning approach based on the polymerase chain reaction, a splice variant of citron, citron kinase has been identified with an alternative amino terminus. This N-terminal extension contains a protein kinase domain that has approximately 50% sequence identity to the sequences of ROCK, ROK, myotonic dystrophy protein kinase and the CDC42 effector known as MRCK or GEK. Citron kinase, which resembles the ROCK family of kinases and by comparison to it, is therefore a multiple domain protein containing an N-terminal kinase domain, an internal coiled-coil domain with Rho/Rac interacting site, and a C-terminal region consisting of a Zn finger, a pleckstrin homology domain, a Citron homology domain, a putative SH3 binding domain, and a PDZ-targeting motif. Its fly ortholog is called Sticky. the importance of different domains of citron-K in its localization at different stages is discussed below.Tissue distribution, localization and dynamics
Investigating the tissue distribution of the citron isoforms with and without the kinase domain, it has been shown that the non-kinase form is restricted to the brain region while the kinase form is widely expressed. Immunofluorescence analyses determined the localization of citron-K and its behavior during cytokinesis. Citron-K first appeared at the equatorial cortex in anaphase, concentrated at the cleavage furrow in early telophase, accumulated in the middle of the intercellular bridge with full ingression of the cleavage furrow in mid telophase, and formed a ring-like structure in the midbody in late telophase, with negligible turnover rate at the midbody. In other words, the protein is much less dynamic at the midbody. Using a series of deletions, it was observed that distinct regions of Citron-K CC domain differentially regulate the localizations of Citron-K during cytokinesis. The C terminal part of the CC domain localized at the cleavage furrow and the midbody while the N-terminal part of the CC domain localized at the central spindle in early telophase and on outer region of the midbody in late telophase.Function
As previously mentioned, citron-K was believed to act in cytokinesis. Its depletion impairs maintenance of the midbody and its overexpression in HeLa cells rendered host cells multinucleated. Cytokinesis failure of Citron-K-depleted cells occurred after full ingression of the cleavage furrow, at the abscission stage. Microtubule disassembly was not seen in any of Citron-K depleted cells with cytokinesis failure. The dominant mode of failure was the inability of daughter cells, which are connected with a shorter intercellular bridge, to separate well. As the midbody microtubules were displaced from the center toward either of the two daughter cells, the two cells fused again with the microtubules absorbed into that daughter cell. To sum the process, Citron-K is important to keep proper structure of the midbody which holds the intercellular bridge microtubules between the two daughter cells and is thus required for successful transition from constriction to abscission.In molecular terms, citron-K depletion impaired the accumulation of 3 key proteins: Rho, Anillin and septins in the intercellular bridge in mid–late telophase, which in previous stages early to mid-telophase was found to co-localize with them.
Interactions
CIT has been shown to interact with RHOB and RHOA.Citron-K or its fly orthologue Sticky has been suggested to interact with several molecules in cytokinesis such as Kinesin-3, actin, myosin light chain, and anillin.