Characterization of the Stat5 Protease

Abstract

Immature myeloid cells have been shown to transduce signals through a carboxyl-terminally truncated isoform of Stat5. This functionally distinct signal transducer and activator of transcription isoform is generated through a unique protein-processing event. Evaluation of numerous cell lines has determined that there is a direct correlation between the expression of truncated Stat5 and protease activity. Moreover, protease activity is found only in the myeloid and not in lymphoid progenitors. To further characterize the protease small quantities have been purified to near homogeneity. Studies on this purified material indicate that the protease has an apparent molecular mass of ϳ25 kDa and is active over a wide range of pH values. The protease will also cleave both activated (i.e. tyrosine-phosphorylated) and inactivate Stat5. Although this activity is sensitive to phenylmethylsulfonyl fluoride, it is notably not sensitive to several other serine protease inhibitors. Additional studies have led to the identification of the unique site where the protease cleaves Stat5. Mutagenesis of this site renders Stat5 resistant to cleavage. Consistent with the model that Stat5 cleavage is important for early myeloid development, introduction of a "non-cleavable" isoform of Stat5 into FDC-P1 cells (a myeloid progenitor line) leads to significant phenotypic changes.

The role of STATs in myeloid differentiation and leukemia

“…This protease cleaves STAT5A and 5B in a perfectly conserved c-terminal motif ATY*MDQA. When this motif was mutated, STAT5 becomes resistant to proteolytic cleavage (Lee et al, 1999). In a preliminary characterization, cleavage-resistant STAT5B partially induced di erentiation of murine FDC-P1, while wildtype STAT5 supported proliferation and self-renewal of these cells.…”

“…In murine cells, p77 and p80 do not result from alternative splicing, but are generated by protein processing. STAT5 is processed by a nuclear serine endopeptidase which is expressed in early murine myeloid, but not lymphoid progenitor cell lines (Azam et al, 1997;Meyer et al, 1998;Lee et al, 1999). This protease cleaves STAT5A and 5B in a perfectly conserved c-terminal motif ATY*MDQA.…”

Signal transduction activator of transcription 5 (STAT5) dysfunction in autoimmune monocytes and macrophages

“…These cytokines can induce signaling through both full-length STAT5A (94-96) and B (94-92) isoforms, as well as through truncated isoforms (77 and 80) that lack the transcriptional activator motif [2,[11][12][13]17,18]. Truncated STAT5 isoforms are not derived from splice variations as seen in other STAT proteins, but are produced post-translationally by the actions of a myeloid-specific nuclear serine protease [13,17].…”

STAT5 isoforms: controversies and clarifications

“…For example, prevalence of this isoform has been reported to be regulated by the maturation state of the cell [22]. Cytokine stimulation also induces the appearance of STAT5 proteolytic isoforms [23]. Piquing interest further is that proteolytically cleaved isoforms have been associated with leukaemia [24,25].…”

“…Through biochemical analysis, Lee et al [23] partially characterized the predominant STAT5 protease in myeloid cells, and found a protein of approx. 25 kDa, but whose identity remained unknown.…”