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Quantitative Evaluation of Alternative Promoter Usage and 3' Splice Variants for Parathyroid Hormone-related Protein by Real-Time Reverse Transcription-PCR

¹ Department of Veterinary Biosciences, College of Veterinary Medicine, and
² Division of Human Cancer Genetics, Department of Molecular Virology, Immunology and Medical Genetics, The Ohio State University, Columbus, OH 43210

^aaddress correspondence to this author at: The Ohio State University, College of Veterinary Medicine, Department of Veterinary Biosciences, 1925 Coffey Rd., Columbus, OH 43210; fax 614-292-6473, e-mail rosol.1@osu.edu

The first 300 words of the full text of this article appear below.

Parathyroid hormone-related protein (PTHrP) was originally isolated from specific cancers as the primary cause of humoral hypercalcemia of malignancy, a paraneoplastic syndrome occurring in humans with a wide variety of malignancies (1). PTHrP also has been reported to be overexpressed by many types of neoplasms not associated with hypercalcemia (2). PTHrP is a polypeptide hormone with structural similarities to parathyroid hormone (PTH) (3)(4). Amino-terminal fragments of PTHrP exert PTH-like actions in bone and kidney by binding to a common receptor for PTH/PTHrP (PTH1 receptor), producing hypercalcemia (5)(6)(7)(8). High expression of PTHrP by cancer cells also has been proposed to play a role in the progression of breast cancer metastasis to bone (9)(10)(11).

The human PTHrP gene is composed of nine exons (Fig. 1A ). Products of exons 5 and 6 are present in all PTHrP transcripts and encode for the prepro region and the majority of the mature peptide. Alternative splicing of the 3' end produces three PTHrP isoforms 139-, 173-, and 141-amino acids in length. Transcriptional regulation of the PTHrP gene is achieved by three distinct promoters located at the 5' end and identified as P1, P2, and P3, respectively. Alternative promoter usage has been evaluated previously by reverse transcription (RT)-PCR based on 5' alternative splicing, and previous studies showed that P3-initiated transcripts were detectable in most tumors, whereas transcripts initiated by either P1 or P2 were present in only a subset of tumors (12)(13)(14)(15)(16).

View larger version (24K): [in this window] [in a new window]   Figure 1. Genomic structure of human PTHrP (A) calibration curve generated by the LightCycler from PTHrP common region calibrators (B), and fluorescence curves for each of the calibrators (C). . . . [Full Text of this Article]

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