Mutations in Promoter Region of Thrombomodulin and Venous Thromboembolic Disease

Abstract

Abstract-The present study was designed to analyze the thrombomodulin proximal promoter region spanning nucleotides Ϫ293 to Ϫ12 to search for polymorphisms that could modify thrombomodulin gene expression in patients with venous thromboembolic disease. The study population comprised 205 patients and 394 healthy subjects of similar age and sex distribution. No polymorphisms and only 1 point mutation (G-33A) were found. The G-33A mutation was present at the heterozygous state in 2 patients and in 1 control. Being more frequent in the patients (0.97%) than in the controls (0.25%), the G-33A mutation might be a risk factor for venous thrombosis. To investigate the effect of this mutation on the thrombomodulin promoter activity, the proximal promoter region of the gene (bearing or not bearing the G-33A mutation) was inserted into a promotorless expression vector, upstream of the firefly luciferase gene, and transiently transfected into EA.hy926 endothelial cells. Under the conditions of the assay, the G-33A mutation mildly decreased the promoter activity. This study confirms that abnormalities of the thrombomodulin proximal promoter are not frequent in patients with venous thromboembolism.

Association of G-33A Polymorphism in the Thrombomodulin Gene with Myocardial Infarction in Koreans.

“…This result basically confirmed the observation by Le Flem et al (12), and seems compatible with the association of 33A with MI; we can expect that, with a lower level of expression of TM, the intravascular environment will tend to be thrombogenic. In contrast to Le Flem et al, however, we found no significant difference in the promoter activity when the shorter constructs were employed.…”

“…This discrepancy might be due to differences in the cell culture systems used in these experiments. In addition, we might have missed small differences in the activity, because the decrease of promoter activity was only 15% in the previous study (12). Our results indicated that the effect of G 33A was at least more obvious in the longer construct than in the shorter one.…”

“…In contrast, Ireland et al found no occurrence of the 33A allele among 168 Caucasians. Le Flem et al confirmed that 33A was rare in Caucasians; they found only 3 heterozygotes among 600 individuals with and without venous thrombosis (12). Therefore, G 33A is much more common in East Asian populations than in Caucasians.…”

Calcium Channel Blockers Act through Nuclear Factor Y to Control Transcription of Key Cardiac Genes

“…Mutations at the CCAAT box, altering it to GTCGA as described previously (Le Flem et al, 1999), were introduced into the full-length TXNIP promoter by fusion PCR using primers 21 to 24 (Supplemental Table S1), and the resulting mutCCAAT-FL construct was verified by sequencing.…”

Naturally occurring mutations in the thrombomodulin gene leading to impaired expression and function

“…20 Each of these latter mutations is in proximity to regulatory elements of the promoter. 21 Only one mutation (G-33A), independently identified in a cohort of patients with venous thrombophilia, was associated with decreased transcriptional activity in reporter gene assay 22 (G.K., unpublished data, August 1999). Furthermore, it was demonstrated that this mutation is associated with CAD in Asian patients.…”