GENOMICS 47, 163–170 (1998) ARTICLE NO. GE975058 Cloning and Characterization of a New Type of Mouse Chemokine Devora L. Rossi,* Gary Hardiman,² Neal G. Copeland,‡ Debra J. Gilbert,‡ Nancy Jenkins,‡ Albert Zlotnik,* and J. Fernando Bazan ² ,1 * Department of Immunobiology and ² Department of Molecular Biology, DNAX Research Institute, Palo Alto, California 94304-1104; and ‡Mammalian Genetics Laboratory, ABL-Basic Research Program, NCI-Frederick Cancer Research and Development Center, Frederick, Maryland 21702 Received June 6, 1997; accepted October 7, 1997 defined by a signature cysteine motif: the a or CXC We report here the identification and characteriza- chemokines, the b or CC chemokines, and the g or C tion of the mouse homologue of a human CX 3 C chemo- chemokine lymphotactin (Kelner et al., 1994; Kennedy kine described by F. Bazan et al. (1997, Nature 385, et al., 1995). These chemokine families chemoattract 640–644). Termed fractalkine, this molecule consti- specific cell populations, and their cognate genes map tutes a fourth or d chemokine structural type that to unique chromosomal locations. The CXC chemokine displays a novel CX 3 C sequence fingerprint. Distinct group induces the migration of neutrophils and lym- from the a, b, or g chemokine families, the polypep- phocytes, while the CC group recruits monocytes, T tide chain of CX 3 C predicts a 373-amino-acid type I cells, basophils, and eosinophils. Moreover, CXC che- transmembrane glycoprotein with the chemokine do- mokines cluster on mouse chromosome 5 (human chro- main resting on top of an extended mucin-like stalk. mosome 4), CC chemokines map to mouse chromosome Comparison of the mouse and human protein chains 11 (human chromosome 17), and the C chemokine shows a high degree of conservation in all the globu- lymphotactin gene resides on mouse and human chro- lar segments with the exception of the stalk portion. mosome 1 (Kennedy et al., 1995). This distinctive fam- The striking identity of an amino acid stretch encom- ily clustering of chemokine genes has a few recent ex- passing a putative juxtamembrane cleavage site sug- ceptions: human MIP-3b has been mapped to chromo- gests the evolutionary conservation of both mem- some 9 (Rossi et al., 1997) and human LARC (also brane-bound and processed CX 3 C forms. Northern named human MIP-3a) to chromosome 2 (Hieshima et analysis reveals the presence of mouse CX 3 C mRNA in al., 1997). heart, brain, lung, kidney, skeletal muscle, and testis We have recently published (Bazan et al., 1997) the tissues. The mouse CX 3 C gene was further localized identification, structure, and in vitro analysis of human to the central region of chromosome 8 by interspecific backcross mapping; a related locus was detected on CX 3 C (or fractalkine). Interestingly, the human CX 3 C chromosome 11. The novel location of this gene from gene was found to reside on human chromosome 16. In other chemokine gene clusters adds to the notion this study we describe the isolation and sequence of that CX 3 C is a fundamentally new class of chemokine. mouse CX 3 C cDNA, characterize the mRNA distribu-  1998 Academic Press tion, and define its (syntenic) chromosomal location. In view of the fundamental role of chemokines in inflammatory diseases and the necessity to under- INTRODUCTION stand their biological function by the use of mouse in vivo models, it was critical to isolate and characterize Chemokines are small proinflammatory peptides, 6– mouse CX 3 C. 14 kDa in size, that direct leukocyte trafficking in im- mune surveillance. Nevertheless, other biological func- MATERIALS AND METHODS tions have been attributed to chemokines including costimulation of cell proliferation, lymphokine produc- Cloning of mouse CX 3 C. Human brain expressed sequence tags tion (Taub et al., 1996), degranulation, cellular activa- (ESTs) that encode mouse CX 3 C were identified in the dbEST data- tion, adhesion, enzyme release, oxidative burst, and base at the National Center for Biotechnology Information by per- forming BLAST searches (Altschul et al., 1990) using the chicken tumor cell cytolysis (Taub et al., 1994, 1995; Lloyd et lymphotactin cDNA (Accession No. AF006742) as the query se- al., 1996). quence. Three of these ESTs were obtained from the I.M.A.G.E. Con- Chemotactic cytokines have been traditionally grouped sortium (Lennon et al., 1996) via Research Genetics (Huntsville, AL) into three structural and functional families that are and sequenced. One of these cDNAs (Accession No. R75309) encoded the chemokine domain, whereas the others corresponded to 3  un- translated sequence (Accession Nos. W74988 and W16003). To clone 1 To whom correspondence should be addressed. Telephone: (415) 496-1115. Fax: (415) 496-1200. E-mail: bazan@dnax.org. the full-length mouse CX 3 C cDNA, oligonucleotides were synthesized 163 0888-7543/98 $25.00 Copyright  1998 by Academic Press All rights of reproduction in any form reserved. AID GENO 5058 / 6r58$$$$41 01-02-98 23:07:30 gnmal