The murine chaperonin 10 gene family contains an intronless, putative genefor early pregnancy factor, Cpn10-rs1

Early pregnancy factor (EPF) is a secreted protein with growth regulatory a nd immunomodulatory properties. Human platelet-derived EPF shares amino aci d sequence identity with chaperonin 10 (Cpn10), a mitochondrial matrix prot ein which functions as a molecular chaperone. The striking differences in c ellular localization and function of the two proteins suggest differential regulation of production reflecting either alternative transcription of the same gene or transcription from different genes. In mammals and more dista ntly related genera, there is a large gene family with homology to CPN 10 c DNA, which includes intronless copies of the coding sequence. To determine whether this could represent the gene for EPF, we have screened a mouse gen omic library and sequenced representative Cpn10 family members, looking for a functional gene distinct from that of Cpn 10, which could encode EPF. Ei ght distinct genes were identified. Cpn10 contains introns, while other mem bers are intronless. Six of these appear to be pseudogenes, and the remaini ng member, Cpn10-rs1, would encode a full-length protein. The 309-bp open r eading frame (ORF) is identical to that of mouse Cpn10 cDNA with the except ion of three single-base changes, two resulting in amino acid changes. Only one further single nucleotide difference between the Cpn10-rs1 and Cpn10 c DNAs is observed, located in the 3' UTR. Single nucleotide primer extension was applied to discriminate between Cpn10-rs1 and Cpn10 expression. Cpn10, which is ubiquitous, was detected in all tissue samples tested, whereas Cp n10-rs1 was expressed selectively. The pattern was completely coincident wi th known patterns of EPF activity, strongly suggesting that Cpn10-rs1 does encode EPF. The complete ORF of Cpn10-rs1 was expressed in E. coli. The pur ified recombinant protein was found to be equipotent with native human plat elet-derived EPF in the bioassay for EPF, the rosette inhibition test.