Intersectin 1 forms a complex with adaptor protein Ruk/CIN85 in vivo independently of epidermal growth factor stimulation

Intersectin multidomain adaptor proteins: Regulation of functional diversity

“…Our recent results demonstrate that ITSN1 forms a stable complex with Ruk/CIN85, a scaffolding protein that regulates recruiting the Cbl-EGFR complex to endosomes for downregulation of RTK, as well as cytoskeletal rearrangements and apoptosis (Nikolaienko et al, 2009a). It should be noted that interactions of the SH3 domains of the ITSN1 with Cbl and Ruk/CIN85 are not regulated by EGF stimulations (Martin et al, 2006;Nikolaienko et al, 2009a,b).…”

Endocytic adaptor protein intersectin 1 forms a complex with microtubule stabilizer STOP in neurons

“…Expression constructs containing GST-fused SH3 domains of ITSN1 were described previously (Nikolaienko et al, 2009). To obtain fluorescent protein (FP)-tagged proteins, full-length coding sequences of STOP and ITSN1-L were amplified by PCR and inserted into pmCherry-N1 (Clontech) and pGFP-C1 (Clontech) vectors, respectively.…”

“…pmCitrine-N1 plasmid was obtained by replacing GFP in pGFP-N1 (Clontech) with mCitrine from mCitrine-SNAP25 construct (obtained from the lab of Dr. Robert Zucker) via digestion with AgeI and BsrGI restriction enzymes and subsequent ligation. Rabbit polyclonal antibodies against the EH2 domain of ITSN1 were described previously (Nikolaienko et al, 2009). Mouse monoclonal anti-STOP antibodies (MAB5524), secondary horseradish peroxidase-labeled (HRP) goat anti-rabbit antibodies and anti-mouse antibodies were purchased from Millipore, Promega and Sigma, respectively.…”

Identification and characterization of a novel mammalian isoform of the endocytic adaptor ITSN1

“…Omni-ITSN1-22aΔ86 was obtained by removing the DNA fragment from the Omni-ITSN1-22a construct via digestion with XhoI and subsequent ligation. The Omni-ITSN1-s construct was described previously (Nikolaienko et al, 2009). The fragment of cDNA encoding SH3A-CTD (residues 745-1025) was subcloned into the pGEX-4T-3 (GE Healthcare) vector.…”

“…Polyclonal antibodies against the EH2 domain of ITSN1 were described previously (Nikolaienko et al, 2009). Anti-Omni (D-8, sc-7270), anti-dynamin 1 (sc-6402) and anti-Cbl (sc-170G) antibodies were purchased from Santa Cruz Biotechnology.…”