Structure of the murine E-selectin ligand 1 (ESL-1) gene and
assignment to Chromosome 8
Frank Willmroth, Arthur L. Beaudet
Department of Human and Molecular Genetics, Baylor College of Medicine, Houston, Texas 77030, USA
Received: 21 April 1999 / Accepted: 12 July 1999
Abstract. A 150-kDa glycoprotein designated in the mouse as
E-selectin ligand-1 (ESL-1; gene symbol Selel) was first isolated
based on its ability to function as a ligand for E-selectin. The gene
appears equivalent to that for membrane glycoprotein MG160 en-
coded in the human by the locus for Golgi apparatus protein 1
(GLG1). ESL-1 is also highly homologous to the chicken cysteine-
rich fibroblast growth factor receptor (CFR). We describe the ge-
nomic structure and chromosomal localization of the Selel locus.
The gene is encoded by 27 exons and extends over approximately
75 kb. It maps to murine Chromosome (Chr) 8 in a region ho-
mologous to human Chr 16q where the GLG1 locus maps, further
indicating that Selel and GLG1 are mouse and human equivalents
of the same gene.
The E-selectin ligand 1 (ESL-1) is expressed on the surface of
various cell lines including fibroblasts, colon carcinoma, and lym-
phoma cells. On neutrophils, ESL-1 is a ligand for E-selectin as
found by affinity chromatography (Steegmaier et al. 1995). E-
selectin is expressed on activated endothelial cells and functionally
involved in early steps of leukocyte adhesion to endothelium
(“tethering”; Kunkel and Ley 1996). In neutrophils, ESL-1 is lo-
cated on microvilli on the cell surface and in the Golgi apparatus
(Steegmaier et al. 1997). The location of adhesion molecules on
leukocyte microvilli is found to be critical in initial contact for-
mation during tethering, most probably owing to the increased
accessibility compared with adhesion molecules which are located
on the planar cell body (von Andrian et al. 1995).
A detailed insight into the molecular interaction of E-selectin
and its ligands is obtained by X-ray crystallography (Graves et al.
1994). In particular, E-selectin residues Glu92, Tyr94, Arg97,
Lys111, and Lys113 have been identified as critical for ligand
binding (Erbe et al. 1992; Graves et al. 1994). The high-affinity
recognition observed between selectins and the carbohydrate-
counter structures on its ligands is based on the binding of multiple
oligosaccharide side chains to the lectin domain of selectin (Nor-
gard et al. 1993; Varki 1994). In the present study, we report the
detailed organization of the murine E-selectin ligand-1 gene (Selel)
and its chromosomal localization. These data provide a basis for
further studies such as the preparation of mutants in the mouse to
study the functions of ESL-1.
Materials and methods
Isolation and characterization of genomic clones.
A murine geno-
mic lambda Fix II phage library (kindly provided by A. Bradley), prepared
from 129/SvEv embryonic stem cells, was used for isolation of genomic
clones except for one clone (lambda clone 3), which originates from a
murine 129/Sv lambda Fix II library (Stratagene, La Jolla, Calif.). Two
1.9-kb PstI and 2-kb NcoI cDNA fragments, derived from the full-length
ESL-1 cDNA kindly provided by D. Vestweber (Steegmaier et al. 1995),
were used for screening of the genomic libraries.
DNA sequence analysis.
Exon-intron boundaries were identified by se-
quencing the exons of the full-length lambda clones with the “PRISM™
Ready Reaction DyeDeoxy™ Terminator Cycle Sequencing Kit” (Applied
Biosytems, Foster City, Calif.). In deviation from the manufacturer’s pro-
tocol, 16 l terminator premix, 1 g template, and 6.4 pmol primer (Gib-
coBRL Life Technologies, Gaithersburg, Md.) were amplified in a 40-l
reaction volume. We ran 25 cycles in a Perkin-Elmer 9600 thermal cycler
(Perkin-Elmer, Norwalk, Conn.) with the following profile: 98°C for 10 s,
50°C for 5 s, 60°C for 4 min. Sequencing was performed with an ABI 377
automated sequencer (Applied Biosystems).
Restriction map construction.
Positive lambda clones were digested
with SacI and XbaI alone and a combination of both. Depending on which
ESL-1 cDNA subfragment hybridized to a particular lambda clone, oligos
representing a part of this region were selected for further hybridizations to
locate the corresponding exon on a lambda clone restriction fragment.
Chromosomal location analysis of the ESL-1 gene (Selel).
blot filters containing PstI-digested genomic DNA isolated from 96 back-
cross animals [(C57BL/6JEi × SPRET/Ei) × SPRET/Ei] were purchased
from The Jackson Laboratory Backcross DNA Panel Mapping Resource
(Rowe et al. 1994) and hybridized with a 227-bp HindIII ESL-1 cDNA
fragment, representing cDNA position 2060–2287 (Acc.# X84037). The
resulting data were sent to The Jackson Lab for pedigree analysis.
Cloning of the E-selectin ligand 1 gene and characterization of
exon/intron boundaries. Approximately 1×10
lambda clones were screened with different subfragments of the
ESL-1 cDNA. Finally, six different overlapping clones were iso-
lated (Table 1, last column). Southern blot hybridization of the
lambda clones with different parts of the ESL-1 cDNA, or oligo-
nucleotides, gave information about the exons coded by each
The murine ESL-1 cDNA, described by Steegmaier et al.
(1995), encodes only 6 bp of 5Ј-untranslated region and is most
probably incomplete at the 5Ј-end, since at least 40 bp are needed
for ribosome scanning and assembly (Kozak 1991, 1996). In com-
parison, the rat homologous MG160 cDNA encodes further up-
stream an open reading frame (uORF), coding 58 amino acids
(Geballe and Morris 1994; Gonatas et al. 1995). Hence, it was of
Database Accession numbers: EMBL/GenBank Y12181 up to Y12206,
Y12462, Mouse Genome Database MGI:104967
Correspondence to: F. Willmroth at Institute for Experimental Oncology,
Am Flughefen 8-10, D-79110 Freiburg, Germany
Mammalian Genome 10, 1085–1088 (1999).
© Springer-Verlag New York Inc. 1999