The 5'-flanking region of the human calreticulin gene shares homology with the human GRP78, GRP94, and protein disulfide isomerase promoters.

R D Sontheimer,J Wilson,D P Mccauliffe,Y S Yang,J D Capra

doi:10.1016/s0021-9258(18)45916-9

R D Sontheimer, J Wilson + Show 3 more

Open Access

https://doi.org/10.1016/s0021-9258(18)45916-9

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Abstract

Calreticulin (CR) is a calcium binding protein that resides in the endoplasmic and sarcoplasmic reticulum and is reactive with human Ro/SS-A autoimmune sera. We have used human CR cDNA to isolate a human 6-kilobase genomic clone that contains 529 base pairs upstream of the presumed transcription start site, 9 exons, 8 introns, and several hundred base pairs 3' of a polyadenylation sequence. Analysis of the human CR promoter region reveals a number of potential regulatory sites also found in the human GRP78, GRP94, and protein disulfide isomerase promoters, including multiple Sp1 and CCAAT consensus sequences, an AP-2 recognition sequence (absent in protein disulfide isomerase), and multiple GC-rich areas. DNA footprint and gel shift analysis on the CR 5'-flanking region demonstrates an area that is bound by protein found in human but not murine nuclear extracts. This sequence is homologous with previously determined regulatory sequences of the human GRP78 and GRP94 promoters. These data indicate that CR, GRP78, GRP94, and protein disulfide isomerase may in part have similar transcriptional regulation and suggest that their gene products while structurally distinct may have similar functions or co-functions. These observations are of additional interest as all four of these genes encode acidic proteins that localize to the endoplasmic reticulum.

Highlights

Calreticulin (CR) is a calcium binding protein that cellular function has not yet been determined
Thissequence is homologous with previously determined regulatory sequences of the human GRP78 and GRP94 promoters
These data indicate that CR, GRP78, GRP94,and proobtained from Dr Richard Baer, University of Texas, Southwestern Medical Center

Summary

Calretieulin exons and introns

3'-most portions of the 1.9-kb CR cDNA. This genomic l ( 1 5 6 ) l(364) I AGzGTAACG,---7/10 TTAG:A insert was excised from the X phage arms in one piece with 2 [105] 2 [184] I AGzGTAAGA,---8/10 TCAG:G. Synthetic oligonucleotides corresponding toCR cDNA sequences of both strands approximate3ly00 base pairs. This is consistent with our earlier Southern filter hybridization datathat showed the gene to be contained and -207 to -211), and several GC-rich areas including four within 6 kb of chromosomal DNA[1].There are exons and putative S p l binding sites The promoter element contains lian genes, most of the introns are type0 or type I (Table I) the AP-2sequence CCCAGGC (-521 to -515) found inSV40. 5"flanking sequence are several putativreegulatory sequences At the 3'-end of the insert is a putative polyadenylation (see Table11) These include a TATA box (-28 to -22), four signal sequence ATTAAA [18].There is a GT-rich area 65-.

GGGCGG CCCAGGC GATTTC TGGTCGACCA CAGCTG GGGNNGGG

DISCUSSION

The multiple CCAAT sequences in thepromoter region are