A splice variant of RILP induces lysosomal clustering independent of dynein recruitment Marije Marsman 1 , Ingrid Jordens 1 , Nuno Rocha, Coenraad Kuijl, Lennert Janssen, Jacques Neefjes * Division of Tumor Biology, The Netherlands Cancer Institute, Amsterdam 1066CX, The Netherlands Received 14 March 2006 Available online 6 April 2006 Abstract The small GTPase Rab7 controls fusion and transport of late endocytic compartments. A critical mediator is the Rab7 effector RILP that recruits the minus-end dynein–dynactin motor complex to these compartments. We identified a natural occurring splice variant of RILP (RILPsv) lacking only 27 amino acids encoded by exon VII. Both variants bind Rab7, prolong its GTP-bound state, and induce clustering of late endocytic compartments. However, RILPsv does not recruit the dynein–dynactin complex, implicating exon VII in motor recruitment. Clustering might still occur via dimerization, since both RILP and RILPsv are able to form hetero- and homo- dimers. Moreover, both effectors compete for Rab7 binding but with different outcome for dynein–dynactin recruitment and transport. Hence, RILPsv provides an extra dimension to the control of vesicle fusion and transport by the small GTPase Rab7. Ó 2006 Elsevier Inc. All rights reserved. Keywords: Rab7; RILP; Late endosomes; Lysosomes; Dynein; MTOC Compartments of the biosynthetic and endocytic path- ways continuously undergo fusion and fission events to maintain their integrity. The family of Rab GTPases regu- lates these processes. Beside regulating membrane fusion, Rab GTPases also regulate membrane transport [1–4]. In principle, one single Rab can regulate distinct steps in these processes by binding to one or more effector proteins. Moreover, each Rab member localizes to a specific com- partment and cycles between an active GTP-bound state and an inactive GDP-bound state, thereby providing a temporal and spatial resolution in the regulation of mem- brane transport and fusion, for review see [4–8]. Rab7 is a key regulator in transport and fusion of late endosomes and lysosomes [9–14]. Three Rab7 effectors have been described, Rab7-interacting lysosomal protein (RILP), oxysterol-binding protein (OSBP)-related pro- tein-1L (ORP1L), and Rabring7 [1,15–17]. Rabring7 has a zinc finger domain and specifically interacts with Rab7- GTP. Expression of Rabring7 results in clustering of late endosomes and lysosomes in the perinuclear area [16]. ORP1L expression can also induce lysosomal clustering. ORP1L has a conserved OSBP-related ligand-binding domain, an N-terminal extension with three ankyrin repeats, and a pleckstrin homology domain (PHD). RILP, which is unrelated to ORP1L and Rabring7, also interacts with Rab7-GTP and overexpression induces clustering of late endosomes and lysosomes [1,15]. In addition, both RILP and ORP1L can induce recruitment of the motor complex dynein–dynactin onto late endosomes and lyso- somes, which facilitates their transport towards the minus-end of the microtubules [1,17]. Recently, the Golgi-restricted Rab34 has been implicat- ed in lysosomal positioning [18]. Overexpression of wild type or constitutively active Rab34 resulted in clustering of late endosomal and lysosomal compartments in the per- inuclear area. Although the precise mechanism has not 0006-291X/$ - see front matter Ó 2006 Elsevier Inc. All rights reserved. doi:10.1016/j.bbrc.2006.03.178 * Corresponding author. Fax: +31 02 5122029. E-mail address: j.neefjes@nki.nl (J. Neefjes). 1 These authors contributed equally to this work. www.elsevier.com/locate/ybbrc Biochemical and Biophysical Research Communications 344 (2006) 747–756 BBRC