The c-Myb oncoprotein is a critical regulator of hematopoietic cell proliferation and differentiation. Normal c-Myb is rapidly degraded by the ubiquitin-26S proteasome pathway, and instability determinants have been localized within the negative regulatory domain in the carboxyl terminus. Our recent work has shown that, in myeloid cells, inhibition of cellular Ser/Thr protein phosphatases with okadaic acid (OA) causes a rapid increase in c-Myb phosphorylation and 26S proteasome-dependent breakdown [J. Bies, S. Feikova, D. P. Bottaro, and L. Wolff (2000) Oncogene 19, 2846-2854]. Furthermore, phosphoamino acid analyses revealed that the increase in phosphorylation was mainly on threonine residues. Here we investigated the ability of c-Myb to bind DNA following phosphorylation. Our results suggest that the hyperphosphorylated form of c-Myb binds to DNA with affinity very similar to the hypophosphorylated form. Therefore, the increased proteolytic instability of the former cannot be explained by a difference in DNA-binding capacity. Conformational changes in the carboxyl terminus were proposed previously to be a consequence of phosphorylation because we observed phosphorylation-induced alterations in gel electrophoresis mobilities and alterations in recognition by specific monoclonal antibodies. Further support for this notion has come from this study, in which we have detected new degradation products in electrophoretic mobility shift assays, as well as an increased rate of in vitro proteolysis, following OA treatment. We speculate that these alterations in the conformation of the negative regulatory domain expose epitopes on the surface of c-Myb, which in turn can serve as recognition signal(s) for ubiquitin-26S proteasome proteolytic machinery.

Amino Acid Sequence↗

PubMedDate not supplied

Three genes with different functions in transformation are regulated by c-Myb in myeloid cells.

The proto-oncogene c-myb is constitutively expressed in murine leukemia virus-induced myeloid leukemia (MML) due to the integration of virus at this locus. Our recent focus has been the determination of genes regulated by this transcription factor that may be involved in transformation. Data presented here, using conditional expression of Myb in myeloid cells, show that c-Myb directly transactivates the endogenous c-myc and Bcl-2 genes, which explains at least in part how c-Myb regulates proliferation and survival. In addition, c-Myb prevents expression at the RNA level of the tumor suppressor INK4b gene. This gene encodes a cyclin-dependent kinase inhibitor, p15INK4b, that is normally upregulated at the mRNA level during myeloid differentiation and promotes growth arrest. The MMLs are generally characterized as differentiated monocytic tumors and possess the phenotype that is normally associated with p15INK4b expression. c-Myb inhibits expression of this gene, however, and therefore acts to promote a pathway which is abnormal in mature cells. This activity of c-Myb collaborates with its maintenance of c-myc expression to promote growth.

Animals↗

PubMedDate not supplied

Toxicological investigations in the semiconductor industry: III: Studies on prenatal toxicity caused by waste products from aluminum plasma etching processes.

The prenatal toxic effects of contaminated vacuum pump oil (Sample A) and solid waste products (Samples B and C) originating from aluminum plasma etching processes in semiconductor manufacturing were investigated. Three strains of pregnant mice with different degrees of sensitivity during organogenesis (days 6-15 of gestation) were treated daily with 1000 mglkg b.w. (Sample A), 500 or 750 mg/kg b.w. (Sample B), and 250, 500, or 750 mg/kg b.w. (Sample C). On day 18 of pregnancy, the animals were killed and examined for gross changes, number and weight of live fetuses, as well as the number of postimplantation losses and malformations. Analytical data showed that the investigated wastes contained various halogenated hydrocarbons and inorganic compounds. Sample A revealed no signs of prenatal toxic action. In contrast, administration of Samples B and C caused strong prenatal toxic effects. The number of live fetuses declined in a dose--related manner, and evidence of intrauterine growth retardation was noted in fetuses that survived to day 18. The number of fetuses with malformations (only cleft palates) rose significantly in accordance with the doses and sensitivity of the strains (> 95 % of the fetuses of the most sensitive strain after 750 mg/kg; 85% after 500 mg/kg).

Aluminum↗