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Functional mutants of the sequence-specific transcription factor p53 and implications for master genes of diversity.

ABSTRACT: There are many sources of genetic diversity, ranging from programmed mutagenesis in antibody genes to random mutagenesis during species evolution or development of cancer. We propose that mutations in DNA sequence-specific transcription factors that target response elements (REs) in many genes can also provide for rapid and broad phenotypic diversity, if the mutations lead to altered binding affinities at individual REs. To test this concept, we examined the in vivo transactivation capacity of wild-type human and murine p53 and 25 partial function mutants. The p53s were expressed in yeast from a rheostatable promoter, and the transactivation capacities toward >15 promoter REs upstream of a reporter gene were measured. Surprisingly, there was wide variation in transactivation by the mutant p53s toward the various REs. This is the first study to address directly the impact of mutations in a sequence-specific transcription factor on transactivation from a wide array of REs. We propose a master gene hypothesis for phenotypic diversity where the master gene is a single transcriptional activator (or repressor) that regulates many genes through different REs. Mutations of the master gene can lead to a variety of simultaneous changes in both the selection of targets and the extent of transcriptional modulation at the individual targets, resulting in a vast number of potential phenotypes that can be created with minimal mutational changes without altering existing protein-protein interactions.

SUBMITTER: Resnick MA

PROVIDER: S-EPMC187891 | biostudies-literature | 2003 Aug

REPOSITORIES: biostudies-literature

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Functional mutants of the sequence-specific transcription factor p53 and implications for master genes of diversity.

Resnick Michael A MA Inga Alberto A

Proceedings of the National Academy of Sciences of the United States of America 20030808 17

There are many sources of genetic diversity, ranging from programmed mutagenesis in antibody genes to random mutagenesis during species evolution or development of cancer. We propose that mutations in DNA sequence-specific transcription factors that target response elements (REs) in many genes can also provide for rapid and broad phenotypic diversity, if the mutations lead to altered binding affinities at individual REs. To test this concept, we examined the in vivo transactivation capacity of w ...[more]

PMID: 12909720

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Project description:The explosion of protein sequences deduced from genetic code has led to both a problem and a potential resource: Efficient data use requires interpreting the functional impact of sequence change without experimentally characterizing each protein variant. Several groups have hypothesized that interpretation could be aided by analyzing the sequences of naturally occurring homologues. To that end, myriad sequence/function analyses have been developed to predict which conserved, semi-conserved, and nonconserved positions are functionally important. These positions must be discriminated from the nonconserved positions that are functionally silent. However, the assumptions that underlie sequence analyses are based on experimental results that are sparse and usually designed to address different questions. Here, we use three homologues from a test family common to bioinformatics-the LacI/GalR transcription repressors-to test a common assumption: If a position is functionally important for one family member, it has similar importance in all homologues. We generated experimental sequence/function information for each nonconserved position in the 18 amino acids that link the DNA-binding and regulatory domains of three LacI/GalR homologues. We find that the functional importance of each position is preserved among the three linkers, albeit to different degrees. We also find that every linker position contributes to function, which has twofold implications. (1) Since the linker positions range from highly conserved to semi-conserved to nonconserved and contribute to affinity, selectivity, and allosteric response, we assert that sequence/function analyses must identify positions in the LacI/GalR linkers to be qualified as "successful". Many analyses overlook this region since most of the residues do not directly contact ligand. (2) No position in the LacI/GalR linker is functionally silent. This finding is inconsistent with another underlying principle of many analyses: Using sequence sets to discriminate important from non-contributing positions obligates silent positions, which denotes that most homologues tolerate a variety of amino acid substitutions at the position without functional change. Instead, additional combinatorial mutants in the LacI/GalR linkers show that particular substitutions can be silent in a context-dependent manner. Thus, specific permutations of sequence change (rather than change at silent positions) would facilitate neutral drift during evolution. Finally, the combinatorial mutants also reveal functional synergy between semi- and nonconserved positions. Such functional relationships would be missed by analyses that rely primarily upon co-evolution.

Dataset Information

Functional mutants of the sequence-specific transcription factor p53 and implications for master genes of diversity.

Publications

Functional mutants of the sequence-specific transcription factor p53 and implications for master genes of diversity.

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