Identification of a novel human tissue factor splice variant that is upregulated in tumor cells

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Abstract
Tissue factor (TF) is a transmembrane glycoprotein that serves as the prime initiator of blood coagulation and plays a critical role in thrombosis and hemostasis. In addition, a variety of tumor cells overexpress cell‐surface TF, which appears to be important for tumor angiogenesis and metastasis. To elucidate the mechanism involved in the upregulation of TF in human tumor cells, a comprehensive analysis of TF mRNA from various normal and tumor cells was performed. The results of these studies indicate that, in addition to possessing a normal full‐length TF transcript and minor levels of an alternatively spliced transcript known as alternatively‐spliced tissue factor (asTF) (Bogdanov et al., Nat Med 2003;9:458–62), human tumor cells express additional full‐length TF transcripts that are also generated by alternative splicing. Reverse transcriptase‐polymerase chain reaction (RT‐PCR) and 5′‐rapid amplification of cDNA ends‐ (5′‐RACE) based analyses of cytoplasmic RNA from normal and tumor cells revealed that there is alternative splicing of the first intron between exon I and exon II resulting in 2 additional TF transcripts. One of the transcripts has an extended exon I with inclusion of most of the first TF intron (955 bp), while the second transcript is formed by the insertion of a 495 bp sequence, referred to as exon IA, derived from an internal sequence of the first intron. The full length TF transcript with alternatively spliced novel exon IA, referred to as alternative exon 1A‐tissue factor (TF‐A), represented ˜1% of the total TF transcripts in normal cells, but constituted 7–10% of the total TF transcript in tumor cells. Quantitative real‐time RT‐PCR analysis indicated that cultured human tumor cells contain 10–25‐fold more copy numbers of TF‐A in comparison to normal, untransformed cells. We propose that high‐level expression of the novel TF‐A transcript, preferentially in tumor cells, may have utility in the diagnosis and staging of a variety of solid tumors.