| dc.description | B2-1 (cytohesin-1) is a member of a group of proteins (including ARNO and ARNO3) that are all of similar size and domain composition. The three proteins contain an N-terminal coiled-coil domain, followed by Sec7 and pleckstrin homology (PH) domains. Recently, several research groups have shown that B2-1 has varied cellular functions and subcellular locations. One of these is an association of the B2-1 Sec7 domain with the plasma membrane, binding to the cytoplasmic portion of the integrin P2 chain (CD 18) and a postulated involvement in inside-out signaling. Other groups have shown that B2-1 and the related proteins are guanine nucleotide-exchange factors that act upon ADP ribosylation factors (ARFs) and are localized to the Golgi or plasma membrane. Here we report the subcellular localization of B2-1 protein. Interestingly, B2-1 does not localize to the plasma membrane, but rather associates with a distinct Golgi complex compartment. B2-1's distribution was disrupted by brefeldin A, a drug that rapidly disrupts the Golgi apparatus by inhibiting ARF activity. Transient transfection of GFP-tagged B2-1 showed Golgi complex targeting. Excessive overexpression of transfected B2-1 also caused partial Golgi dispersion. While it is well established that the Sec7 domain has GEF activity and the PH domain anchors the proteins to membrane phosphoinositols, the function of the N-terminal coiled-coil region is unknown. Here it was shown that B2-1's N-terminus (residues 1--54) is necessary and sufficient to target the protein to the Golgi. The Sec7+PH domains of B2-1 (residues 55--398) were not sufficient for Golgi localization. Further deletion analysis and point mutagenesis indicated that the coiled-coil domain within the N terminus is responsible for Golgi targeting. Furthermore, ARNO and ARNO3 N termini also have the same capability for targeting to the Golgi. It was concluded that the N-terminal a-helical coiled-coil domain is used to target this family of proteins to the Golgi complex. | en_US |
