No abstract

The oncogene v‐myb and its cellular progenitor c‐myb encode nuclear, DNA binding phosphoproteins that control the expression of certain target genes in immature hematopoietic cells. Here, we report the isolation of a myb‐related chicken gene, chicken B‐myb. We show that expression of B‐myb, unlike that of c‐myb, is not restricted to hematopoietic cells, suggesting that B‐myb functions in a broader spectrum of cell types than c‐myb. We have identified the authentic chicken B‐myb protein as a nuclear protein of approximately 110 kDa. We show that the B‐myb protein specifically recognizes v‐myb binding sites in vitro and that binding is mediated by an N‐terminally located DNA binding domain. Although B‐myb protein recognizes myb binding sites, B‐myb fails to transactivate several myb‐responsive gene constructs as well as the endogenous myb‐responsive gene mim‐1. Instead, we find that B‐myb represses v‐myb‐ and c‐myb‐mediated activation of the mim‐1 gene, most likely by competing with other myb proteins for binding sites. Our results raise the possibility that B‐myb is an inhibitory member of the myb family.

No abstract

Antisera against vertebrate calcium-binding proteins cross-react with Drosophila nervous and muscle tissue. We have used an antiserum against carp parvalbumin to isolate from a Drosophila head cDNA library immunopositive expression clones. Tissue in situ hybridization identified a clone that labeled specific neurons and muscles similar to the parvalbumin-like immunohistochemical staining pattern. Five independent cDNAs derive from an mRNA whose open reading frame codes for a 310 amino acid polypeptide. Sequence analysis identifies six EF-hand calcium-binding domains and reveals 42% and 37% homology to chicken calretinin and calbindin D-28k, respectively. Since the positions of 9 out of 10 introns within the ORF are conserved from the Drosophila gene to both vertebrate genes, we conclude that we have identified the first invertebrate member of the calbindin sub-family of calcium-binding protein genes of the EF-hand homolog family. The calbindin-32 gene (cbn) maps to 53E on the second chromosome. It is expressed through most of ontogenesis with a selective distribution in the nervous system and in a few small adult thoracic muscles. The cloning of a Drosophila homolog to vertebrate neuronal Ca(2+)-binding proteins opens new routes to study the so far largely elusive function of these brain molecules.

We isolated five monoclonal antibodies (mAbs) made against tetracycline repressor (TetR), one against the TetR tetracycline complex (Tc) and two against theTetR‐tet operator (tetO) complex. The epitopes of the anti‐TetR mAbs are localized in the α‐helix‐turn‐α‐helix motif (HTH), at different sites near the Tc binding pocket and at the dimerization interface. The anti‐TetR‐Tc and one of the anti‐TetR‐tetO mAbs recognize epitopes near the Tc binding pocket. The other anti‐TetR‐tetO mAb binds to an epitope within the HTH. Quantitative immunoprecipitation and competitive ELISA employing TetR, TetR‐Tc, or TetR‐tetO revealed different affinities of the mAbs for TetR in these functional states. Binding of the two mAbs to epitopes in the HTH was identical for TetR and TetR‐Tc indicating the same conformation in both forms. The epitope located in the dimerization interface is bound more strongly in TetR compared to TetR‐Tc, supporting the idea of different conformations of that epitope in these forms of TetR. The greatest affinity differences were found for epitopes around the Tc binding pocket. Two anti‐TetR mAbs have the highest affinities for free TetR, somewhat reduced affinity for TetR‐tetO and the lowest affinities for TetR‐Tc. The anti‐TetR‐Tc mAb has a discontinuous epitope, formed in TetR‐Tc, which is less well bound in TetR and not bound in the TetR‐tetO complex. One anti‐TetR‐tetO mAb does not recognize TetR‐Tc. Since the epitopes do not overlap with the respective ligand binding sites on TetR, these results are interpreted as conformational differences of the epitopes in these forms of TetR.