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L Kretzner

Publications and source records attributed to L Kretzner.

10 recordsLinked to original sources
PubMed1992

Myc and Max proteins possess distinct transcriptional activities.

The Myc family proteins are thought to be involved in transcription because they have both a carboxy-terminal basic-helix-loop-helix-zipper (bHLH-Z) domain, common to a large class of transcription factors, and an amino-terminal fragment which, for c-Myc, has transactivating function when assayed in chimaeric constructs. In addition, c-, N- and L-Myc proteins heterodimerize, in vitro and in vivo, with the bHLH-Z protein Max. In vitro, Max homodimerizes but preferentially associates with Myc, which homodimerizes poorly. Furthermore Myc-Max heterodimers specifically bind the nucleotide sequence CACGTG with higher affinity than either homodimer alone. The identification of Max and the specific DNA-binding activities of Myc and Max provides an opportunity for directly testing the transcriptional activities of these proteins in mammalian cells. We report here that Myc overexpression activates, whereas Max overexpression represses, transcription of a reporter gene. Max-induced repression is relieved by overexpression of c-Myc. Repression requires the DNA-binding domain of Max, whereas relief of repression requires the dimerization and transcriptional activation activities of Myc. Both effects require Myc-Max-binding sites in the reporter gene.

Animals↗
PubMed1992

Transcriptional activities of the Myc and Max proteins in mammalian cells.

The myc family of oncogenes exhibit deregulated expression in a host of neoplasias. Though the molecular function of the Myc protein in both normal and tumorigenic cells has remained uncertain, it has been postulated to play a role in gene transcription on the basis of amino acid homologies with known transcription factors such as MyoD (Lüscher & Eisenman, 1990). We report here the direct testing of full-length Myc and its dimerization partner, Max, on the transcriptional activity of reporter genes bearing Myc/Max binding sites. Such reporter constructs display an endogenous level of activity in transient transfections which is dependent on the presence of the CACGTG sequence. Exogenous expression of myc results in modest activation of reporter gene transcription. Similar overexpression of max results in a repression of reporter gene activity, an effect which is reversed by co-expression with c-myc. Max repression is dependent on an intact DNA binding region, while Myc activation depends on both the N-terminal activation and the C-terminal dimerization domains. These results suggest a model in which Max homodimers can act as as repressors, and Myc-Max heterodimers as activators, of potential target genes.

Base Sequence↗
PubMed1992

Myc and Max function as a nucleoprotein complex.

The Myc family of oncoproteins are thought to regulate proliferation and differentiation in a wide variety of cell types. Recent studies show that Myc proteins form sequence-specific DNA-binding complexes with Max, a new member of the helix-loop-helix leucine zipper protein class. The properties of the Myc-Max complex suggest a mechanism for Myc's function in both normal and neoplastic cell behavior.

Basic-Leucine Zipper Transcription Factors↗
PubMed1990

Sequence-specific DNA binding by the c-Myc protein.

While it has been known for some time that the c-Myc protein binds to random DNA sequences, no sequence-specific binding activity has been detected. At its carboxyl terminus, c-Myc contains a basic--helix-loop-helix (bHLH) motif, which is important for dimerization and specific DNA binding, as demonstrated for other bHLH protein family members. Of those studied, most bHLH proteins bind to sites that contain a CA- -TG consensus. In this study, the technique of selected and amplified binding-sequence (SAAB) imprinting was used to identify a DNA sequence that was recognized by c-Myc. A purified carboxyl-terminal fragment of human c-Myc that contained the bHLH domain bound in vitro in a sequence-specific manner to the sequence, CACGTG. These results suggest that some of the biological functions of Myc family proteins are accomplished by sequence-specific DNA binding that is mediated by the carboxyl-terminal region of the protein.

Base Sequence↗
PubMed1990

Saccharomyces cerevisiae U1 small nuclear RNA secondary structure contains both universal and yeast-specific domains.

The five small nuclear RNAs (snRNAs) involved in mammalian pre-mRNA splicing (U1, U2, U4, U5, and U6) are well conserved in length, sequence, and especially secondary structure. These five snRNAs from Saccharomyces cerevisiae show notable size and sequence differences from their metazoan counterparts. This is most striking for the large S. cerevisiae U1 and U2 snRNAs, for which no secondary structure models currently exist. Because of the importance of U1 snRNA in the early steps of "spliceosome" assembly, we wanted to compare the highly conserved secondary structure of metazoan U1 snRNA (approximately 165 nucleotides) with that of S. cerevisiae U1 snRNA (568 nucleotides). To this end, we have cloned and sequenced the U1 gene from two other yeast species possessing large U1 RNAs. Using computer-derived structure predictions, phylogenetic comparisons, and structure probing, we have arrived at a secondary structure model for S. cerevisiae U1 snRNA. The results show that most elements of higher eukaryotic U1 snRNA secondary structure are conserved in S. cerevisiae. The hundreds of "extra" nucleotides of yeast U1 RNA, also highly structured, suggest that large insertions and/or deletions have occurred during the evolution of the U1 gene.

Base Sequence↗
PubMed1990

Universally conserved and yeast-specific U1 snRNA sequences are important but not essential for U1 snRNP function.

To study the contribution of the large, 568-nucleotide yeast (Saccharomyces cerevisiae) U1 snRNA to pre-mRNA splicing, we generated mutations in two regions of the molecule and introduced each mutant gene back into yeast as the sole copy of the U1 snRNA gene. We mutagenized the "A loop," a subregion highly conserved in primary sequence in all U1 snRNA molecules analyzed to date. We also mutagenized a portion of the yeast core subdomain, a region conserved in primary and secondary structure among several yeast species but absent from the much smaller metazoan U1 molecule. Surprisingly, mutations in these two regions had little or no effect on growth rate, yet several of them affected an inefficiently spliced reporter gene construct. In addition, combinations of mutants in both regions gave rise to reduced growth rates. Using the latter assay, we confirmed some of the proposed secondary structure of the yeast core domain. The experiments indicate that both regions contribute to U1 snRNP activity but that mutations in a single region do not have a substantial effect on growth rate because U1 snRNP activity is not rate-limiting for growth.

Base Sequence↗
PubMed1988

A U1 snRNA:pre-mRNA base pairing interaction is required early in yeast spliceosome assembly but does not uniquely define the 5' cleavage site.

We analyzed the effects of suppressor mutations in the U1 snRNA (SNR19) gene from Saccharomyces cerevisiae on the splicing of mutant pre-mRNA substrates. The results indicate that pairing between U1 snRNA and the highly conserved position 5 (GTATGT) of the intron occurs early in spliceosome assembly in vitro. This pairing is important for efficient splicing both in vitro and in vivo. However, pairing at position 5 does not appear to influence 5' splice site selection in vivo, indicating that the previously described U1 snRNA:5' splice junction base pairing interaction is not sufficient to define the 5' cleavage site.

Introns↗
PubMed1987

S. cerevisiae U1 RNA is large and has limited primary sequence homology to metazoan U1 snRNA.

We have cloned and sequenced the yeast SNR19 gene and show here that snR19 is the yeast homolog of metazoan U1 snRNA. sn R19 is 569 nucleotides long, strikingly larger than its metazoan counterpart. The two molecules resemble each other closely in the predicted secondary structure of their first 50 nucleotides. Primary sequence homology is restricted to some of their single-stranded regions, including 11 consecutive nucleotides at the 5' end of the two molecules, the region that interacts with pre-mRNA 5' splice junctions. snR19 is spliceosome-associated and required for in vitro pre-mRNA splicing. We also note that 8 sequences in snR19 have extensive complementarity to snR20, the large yeast U2 RNA, suggesting that yeast U1 may interact with yeast U2 by base-pairing.

Animal Population Groups↗