No abstract

Stat proteins (signal transducers and activators of transcription) are transcription factors which transmit signals from activated cytokine receptors to the nucleus. They are tyrosine phosphorylated by Jak kinases and dimerize through their SH2 domains (22,59). A functional domain structure of the Stat proteins has been established, and the regions responsible for dimerization, specific DNA binding, and transactivation have been identified (21,23,49,62). Stat5a, Stat5b, and Stat6 are the members of the Stat gene family (26) which have the highest amino acid homologies. These mammalian Stat factors are related to Marelle, a recently identified Stat protein important for Drosophila larval development (25, 76). The sequence homologies of Marelle with Stat5 and Stat6 are 37 and 34%, respectively. Stat5a and Stat5b are two highly related genes expressed in most tissues (3,31,41,51

Proteasomal proteolysis relies on the activity of six catalytically active proteasomal subunits (b1, b2, b5, b1i, b2i and b5i). Applying a functional proteomics approach, we used a recently developed activity-based, cell-permeable proteasome-specific probe that for the first time allows differential visualization of individual active proteasomal subunits in intact primary cells. In primary leukemia samples, we observed remarkable variability in the amounts of active b1/1i-, b2/2i-and b5/5i-type of subunits, contrasting with their constant protein expression. Bortezomib inhibited b5-and b1-type, but to a lesser extend b2-type of subunits in live primary cells in vitro and in vivo. When we adapted the bortezomib-sensitive human acute myeloid leukemia cell line HL-60 to bortezomib 40 nM (HL-60a), proteasomal activity profiling revealed an upregulation of active subunits, and residual b1/b5-type of activity could be visualized in the presence of bortezomib 20 nM, in contrast to control cells. In a panel of cell lines from hematologic malignancies, the ratio between b2-type and (b1 þ b5)-type of active proteasomal polypeptides mirrored different degrees of bortezomib sensitivity. We thus conclude that the proteasomal activity profile varies in primary leukemia cells, and that the pattern of proteasomal subunit activity influences the sensitivity of hematologic malignancies toward bortezomib. Leukemia (2007) 21, 84-92.

Inhibition of BCR-ABL

The possible role of homodimerization events in intracellular signal transduction triggered by the bipartite human interleukin-4 receptor was addressed. We generated cell lines functionally expressing derivatives of the two receptor subunits alpha and gamma, which allow for a specific and background-free experimental induction of intracellular homo- and heterodimers. A heterodimer of alpha and gamma released an intracellular signal, whereas a gamma-gamma homodimer did not. Unexpectedly, we found the intracellular domain of interleukin-4 receptor alpha chain to evoke cell proliferation and activation of tyrosine kinase Jak1 as well as of transcription factor Stat6 upon homodimerization. Both recruitment of the common gamma chain and activation of kinase Jak3 were shown to be dispensible for these processes.