A functional SUMO-interacting motif in the transactivation domain of c-Myb regulates its myeloid transforming ability

c-Myb is an essential hematopoietic transcription factor that controls proliferation and differentiation of progenitors during blood cell development. Whereas sumoylation of the C-terminal regulatory domain (CRD) is known to have a major impact on the activity of c-Myb, no role for noncovalent binding of small ubiquitin-like modifier (SUMO) to c-Myb has been described. Based on the consensus SUMO-interacting motif (SIM), we identified and examined putative SIMs in human c-Myb. Interaction and reporter assays showed that the SIM in the in the transactivation domain of c-Myb (V 267 NIV) is functional. This motif is necessary for c-Myb to be able to interact noncovalently with SUMO, preferentially SUMO2/3. Destroying the SUMO-binding properties by mutation resulted in a large increase in the transactivation potential of c-Myb. Mutational analysis and overexpression of conjugation-defective SUMO argued against intramolecular repression caused by sumoylated CRD and in favor of SUMO-dependent repression in trans. Using both a myeloid cell line-based assay and a primary hematopoietic cell assay, we addressed the transforming abilities of SUMO binding and conjugation mutants. Interestingly, only loss of SUMO binding, and not SUMO conjugation, enhanced the myeloid transformational potential of c-Myb. c-Myb with the SIM mutated conferred a higher proliferative ability than the wild-type and caused an effective differentiation block. This establishes SUMO binding as a mechanism involved in modulating the transactivation activity of c-Myb, and responsible for keeping the transforming potential of the oncoprotein in check

The Myb leucine zipper is essential for leukemogenicity of the v-Myb protein

The AMV v-Myb oncoprotein causes oncogenic transformation of myelomonocytic cells in vivo and in vitro. Its transforming capacity is strictly dependent upon the N-terminal DNA binding domain, the central transactivation region, and on the C-terminal domain containing a putative leucine zipper motif. Here we show that the v-MybL3,4A mutant, in which Leu325 and Leu332 of the leucine zipper have been replaced by alanines, failed to induce leukemia in virus infected chicken. This demonstrates that the leucine zipper domain is indispensable for v-myb induced leukemogenesis in vivo. v-MybL3,4A was, however, still able to transform myelomonocytic cells from chicken bone marrow in vitro. Yet, while v-mybL3,4A transformed cells were impaired in growth at 37 degrees C, they failed to grow at 42 degrees C, the physiological body temperature of avian species. This might explain the loss of v-MybL3,4A leukemogenic potential in vivo. We also demonstrate that the v-Myb leucine zipper domain interacts in vitro with two host cell proteins, p26 and p28. This interaction is compromised in v-MybL3,4A indicating that binding of v-Myb to p26 and p28 might be important for the leukemogenic potential of v-Myb

bFGF signaling and v-Myb cooperate in sustained growth of primitive erythroid progenitors

The development of red blood cells from hematopoietic progenitors requires the interplay of specific extracellular factors and transcriptional regulators. Here we have identified an erythroid progenitor that is critically dependent on bFGF and requires expression of AMV v-Myb for sustained proliferation in vitro, indicating that bFGF and Myb proteins cooperate in these cells. In the presence of bFGF such v-Myb cells are completely blocked in their ability to differentiate and exhibit an exceptionally high proliferative potential and long life-span in vitro. Interestingly, in the absence of bFGF cells effectively differentiate into mature erythrocytes, irrespective of constitutive and elevated levels of v-Myb. We also demonstrate that these cells express high levels of FGF receptor type 1 (FGFR1) and that phospholipase C{gamma} (PLCT{gamma}) is one of the important molecules in FGF receptor signaling. Our studies suggest that bFGF, in cooperation with Myb proteins, represents an important factor for determining erythroid lineage choice. These findings unravel a so far unidentified link between extracellular signaling and Myb in hematopoietic cells

The leucine zipper region of Myb oncoprotein regulates the commitment of hematopoietic progenitors

The development of blood cells proceeds from pluripotent stem cells through multipotent progenitors into mature elements belonging to at least 8 different lineages. The lineage choice process during which stem cells and progenitors commit to a particular lineage is regulated by a coordinated action of extracellular signals and transcription factors. Molecular mechanisms controlling commitment are largely unknown. Here, the transcription factor v-Myb and its leucine zipper region (LZR) are identified as regulators of the commitment of a common myeloid progenitor and progenitors restricted to the myeloid lineage. It is demonstrated that wild-type v-Myb with the intact LZR directs development of progenitors into the macrophage lineage. Mutations in this region compromise commitment toward myeloid cells and cause v-Myb to also support the development of erythroid cells, thrombocytes, and granulocytes, similar to the c-Myb protein. In agreement with that, the wild-type v-Myb induces high expression of myeloid factors C/EBPβ, PU.1, and Egr-1 in its target cells, whereas SCL, GATA-1, and c-Myb are more abundant in cells expressing the v-Myb LZR mutant. It is proposed that Myb LZR can function as a molecular switch, affecting expression of lineage-specifying transcription factors and directing the development of hematopoietic progenitors into either myeloid or erythroid lineages

Transcribing meetings with the AMIDA systems

In this paper, we give an overview of the AMIDA systems for transcription of conference and lecture room meetings. The systems were developed for participation in the Rich Transcription evaluations conducted by the National Institute for Standards and Technology in the years 2007 and 2009 and can process close talking and far field microphone recordings. The paper first discusses fundamental properties of meeting data with special focus on the AMI/AMIDA corpora. This is followed by a description and analysis of improved processing and modeling, with focus on techniques specifically addressing meeting transcription issues such as multi-room recordings or domain variability. In 2007 and 2009, two different strategies of systems building were followed. While in 2007 we used our traditional style system design based on cross adaptation, the 2009 systems were constructed semi-automatically, supported by improved decoders and a new method for system representation. Overall these changes gave a 6%-13% relative reduction in word error rate compared to our 2007 results while at the same time requiring less training material and reducing the real-time factor by five times. The meeting transcription systems are available at www.webasr.org