Tong J H, Fant X, Benoit G, Chen S J, Chen Z, Lanotte M
Centre G. Hayem, I.N.S.E.R.M. U-496, Hôpital Saint-Louis, 1, Avenue Claude Vellefaux, Paris, 75010, France.
Genomics. 2000 Nov 1;69(3):380-90. doi: 10.1006/geno.2000.6347.
The Jem-1 (JEM-1, HGMW-approved symbol BLZF1) gene mapping to human chromosome 1q24 codes for a ubiquitously expressed 3-kb mRNA, translated in a 45-kDa nuclear protein. Recent studies have shown a deficient expression of this gene in acute promyelocytic leukemia (APL). However, treatment with retinoids was able to upregulate JEM-1 mRNA in maturing NB4 leukemia cells. Here, we report the characterization of the structural organization of JEM-1. By hybridization screening of a human genomic library derived from blood mononuclear cells, five overlapping genomic DNA clones were isolated. These clones extend over 34 kb of the human genome and comprise the complete JEM-1 gene and a 4-kb 5'flanking region. Determination of the exon-intron structure of Jem-1 revealed seven exons whose junctions with introns exhibited typical splice sequences. A shorter transcript (Jem-1s, 1.3 kb) generated by exon 3 extension and polyadenylation was identified. Its translation generated a 23-kDa protein that exhibited a cytoplasmic localization. 5'RACE-PCR identified a major transcription start site (TSS) located at 403 nt upstream of the ATG. Computer analysis of the 1. 8-kb 5'flanking region showed that it lacks a TATA box, Inr motifs or DPE motifs, but it contains a typical CCAAT box located 95 bp upstream of the TSS. Sequencing also revealed potential cis-acting elements for multiple transcription regulators including Sp1, GATA, C/EBP, AP-1, and Pu1. No retinoic acid receptor elements or retinoic X receptor elements were detected. This 1.8-kb DNA sequence showed a strong constitutive promoter activity determined by a luciferase-reporter gene assay in transiently transfected HeLa cells. Retinoids further increased luciferase expression 2.7-fold. We demonstrated that the 1-kb distal sequence contains yet unidentified elements reducing constitutive transcription. Thus, the maximal constitutive promoter activity was assigned to a -432 + 101 region overlapping the TSS. These data support the idea of a constitutive expression of JEM-1, but a negative regulation in APL released by retinoids.
定位于人类1号染色体1q24的Jem-1（JEM-1，HGMW认可符号BLZF1）基因编码一个广泛表达的3kb mRNA，其翻译产物为一种45kDa的核蛋白。最近的研究表明，该基因在急性早幼粒细胞白血病（APL）中表达不足。然而，用维甲酸处理能够使成熟的NB4白血病细胞中的JEM-1 mRNA上调。在此，我们报告JEM-1结构组织的特征。通过对来自血液单核细胞的人类基因组文库进行杂交筛选，分离出了五个重叠的基因组DNA克隆。这些克隆覆盖了人类基因组的34kb，包含完整的JEM-1基因和一个4kb的5'侧翼区域。Jem-1外显子-内含子结构的测定揭示了七个外显子，其与内含子的连接处呈现典型的剪接序列。鉴定出了一个由外显子3延伸和多聚腺苷酸化产生的较短转录本（Jem-1s，1.3kb）。其翻译产生了一种定位于细胞质的23kDa蛋白。5'RACE-PCR确定了一个主要转录起始位点（TSS），位于ATG上游403nt处。对1.8kb的5'侧翼区域进行计算机分析表明，它缺乏TATA盒、Inr基序或DPE基序，但在TSS上游95bp处含有一个典型的CCAAT盒。测序还揭示了多个转录调节因子的潜在顺式作用元件，包括Sp1、GATA、C/EBP、AP-1和Pu1。未检测到维甲酸受体元件或维甲酸X受体元件。通过荧光素酶报告基因检测，在瞬时转染的HeLa细胞中测定该1.8kb DNA序列显示出很强的组成型启动子活性。维甲酸进一步使荧光素酶表达增加2.7倍。我们证明1kb的远端序列包含尚未鉴定的降低组成型转录的元件。因此，最大组成型启动子活性被指定给与TSS重叠的-432 + 101区域。这些数据支持JEM-1组成型表达的观点，但在APL中由维甲酸释放负调控。
