Human eosinophil Charcot-Leyden crystal protein: cloning and characterization of a lysophospholipase gene promoter.
The Charcot-Leyden crystal (CLC) protein is a lysophospholipase expressed exclusively by eosinophils and basophils. During eosinophilic differentiation of eosinophil-committed cell lines, CLC steady state mRNA levels increase significantly. This increased expression is transcriptionally regulated during butyrate induction of an eosinophilic subline (C15) of the promyelocytic leukemia cell line HL-60, as shown by nuclear run-on assays. The transcriptional start site of the CLC gene was identified 43 bp upstream of the 5' end of the longest available cDNA sequence. The gene encoding CLC protein was cloned from a chromosome 19-specific library and a fragment overlapping the transcriptional start site was isolated and sequenced. Plasmid constructs (in the pXP2 luciferase expression vector) containing 411 and 292 bp of genomic sequence upstream of the CLC transcriptional start site directed reporter gene expression in transient transfections of HL-60-C15 cells, as well as other myeloid (U937) and nonmyeloid (HeLa and RPMI 8402) cell lines. However, the differential expression of the two CLC promoter constructs in these cell lines suggests that the -292 to -411 bp region of the promoter may confer some specificity for expression in the eosinophil lineage. The CLC promoter sequence contains two consensus GATA binding sites, a purine-rich sequence that presents potential binding sites for PU.1, a member of the ets family of genes, as well as sequences described in other myeloid-specific promoters. This is the first demonstration of a functional eosinophil promoter that could serve as a model for identifying DNA elements and trans-activating factors that regulate gene expression during the commitment and differentiation of the eosinophil lineage.
Gomolin
H I
HI
Department of Medicine, Beth Israel Hospital, Boston, MA 02215.
Yamaguchi
Y
Y
Paulpillai
A V
AV
Dvorak
L A
LA
Ackerman
S J
SJ
Tenen
D G
DG
D000595:Amino Acid Sequence; D001483:Base Sequence; D015152:Blotting, Northern; D002467:Cell Nucleus / Q000378:metabolism; D002888:Chromosomes, Human, Pair 19; D003001:Cloning, Molecular; D004804:Eosinophils / Q000201:enzymology*; D015698:Genomic Library; D006023:Glycoproteins / Q000096:biosynthesis* / Q000235:genetics; D006801:Humans; D007438:Introns; D015473:Leukemia, Promyelocytic, Acute; D008245:Lysophospholipase / Q000235:genetics*; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000096:biosynthesis / Q000097:blood / Q000302:isolation & purification; D015183:Restriction Mapping; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured
D006023:Glycoproteins; D015345:Oligonucleotide Probes; D012333:RNA, Messenger; D008245:Lysophospholipase; C011711:lysolecithin acylhydrolase