A complementary DNA (2510 bp) encoding a protein with high homology to mouse phospholipase A2-activating protein (PLAP) was cloned from rat hepatocytes by differential screening of a subtractive (normoxic minus hypoxie) λGEM-2 cDNA library. This cDNA clone, denoted as HCW9, encodes a 647-amino-acid (aa) protein with 96.1% identity in the N-terminal portion (255 aa) to mouse PLAP [Clark et al., Proc. Natl. Acad. Sci. USA 88 (1991) 5418-5422]. Four cDNA clones were further isolated by screening a primary normoxic λGEM-2 cDNA library with a fragment of the HCW9 clone as probe. Partial sequencing of these clones revealed that all of the four clones were identical to clone HCW9. Northern blotting analysis of total RNA extracted from normoxic or hypoxic rat hepatocytes, with two different fragments of clone HCW9 as probes, demonstrated a single band with a mobility corresponding to a size of 2.5 kb, whose level was significantly decreased during chemical hypoxia.
Bibliographical noteFunding Information:
We thank Dr. Mike A. Clark of the Smith Kline and French Laboratories (King of Prussia, PA, USA) for his generous gift of the mouse PLAP clone. We also thank Dr. Wayne Litaker of the Biotechnology Center of University of North Carolina for his valuable discussions and CD. Harrsion for her technical assistance during the course of this research. This research was supported by grant AGO7218 from NIH to B.H.
- amino-acid sequence
- bee venom melittin
- chemical hypoxia
- nucleotide sequence
- phage library
- recombinant DNA