PURIFICATION AND CDNA CLONING OF SAPKK3, THE MAJOR ACTIVATOR OF RK P38 IN STRESS-STIMULATED AND CYTOKINE-STIMULATED MONOCYTES AND EPITHELIAL-CELLS/

Two chromatographically distinct stress-activated protein kinase kinas es (SAPKKs) have been identified in several mammalian cells, termed SA PKK2 and SAPKK3, which activate the MAP kinase family member RK/p38 bu t not JNK/SAPK in vitro. Here we demonstrate that SAPKK2 is identical or very closely related to the MAP kinase kinase family member MKK3, H owever under our assay conditions, SAPKK3 was the major activator of R K/p38 detected in extracts prepared from stress- or interleukin-1-stim ulated epithelial (KB) cells, from bacterial lipopolysaccharide- and t umour necrosis factor alpha-stimulated THP1 monocytes or from rabbit s keletal muscle, The activated form of SAPKK3 was purified from muscle to near homogeneity, and tryptic peptide sequences were used to clone human and murine cDNAs encoding this enzyme. Human SAPKK3 comprised 33 4 amino acids and was 78% identical to MKK3, The marine and human SAPK KS were 97% identical in their amino acid sequences, We also cloned a different murine cDNA that appears to encode a SAPKK3 protein truncate d at the N-terminus, SAPKK3 is identical to the recently cloned MKK6.