MOLECULAR BASIS OF TRANSLATIONAL RECODING IN YEAST
MOLECULAR BASIS OF TRANSLATIONAL RECODING IN YEAST
批准号:
2900549
负责人:
Philip James Farabaugh
金额:
$30.03万
依托单位国家:
美国
项目类别:
财政年份:
1989
资助国家:
美国
项目状态:
已结题
起止时间:
1989-08-01 至 2002-03-31
关键词:
RNA splicing Saccharomyces endonuclease frameshift mutation fungal genetics gel electrophoresis gene expression genetic mapping genetic promoter element genetic recombination genetic transcription genetic translation mass spectrometry molecular cloning nucleic acid hybridization nucleic acid sequence oligonucleotides polymerase chain reaction ribosomes site directed mutagenesis transposon /insertion element
中文摘要
该建议涉及+1翻译移码的机制,
这两种事件都发生在特殊的mRNA位点,
阅读框架(程序性移码)和表型抑制
由突变型tRNA引起的移码突变(移码抑制)。
长期以来人们一直认为这两件事没有关联
机械地。 移码抑制的经典模型
提出了一种具有扩展的反密码子环的突变tRNA,
扩展了四个核苷酸的反密码子,导致下游的转移,
或者+1方向。 相比之下,程序化的移码现在似乎发生了
因为最后一个框内密码子的近同源解码,
到下一个密码子移位阅读时的翻译错误。 的
经典的抑制模型现在看来是无效的,抑制可能
通过类似于编程移码的机制发生。 这项建议
寻求进一步探索程序化框架转换的机制,
为了测试用于移码的近同源解码模型是否
压制是正确的。
酵母分子遗传学分析将用于进一步测试
近同源解码模型的有效性
移码 基于PCR的基因破坏将消除每一个
酵母基因组中的低拷贝tRNA基因;在这些菌株中,
密码子必须由近同源的同种受体读取,这可能诱导高表达。
效率框架转换 在某些情况下,发生移码是因为
正常的同源tRNA不能有效地解码doo;位点特异性
诱变将鉴定tRNA中有助于
他们的低效率。 一种促进+1的体外翻译系统
编程移码将用于识别哪些近同源
tRNA实际上负责促进框架的转变。
最后,顺式作用mRNA的移码刺激的基础
序列,并通过延伸因子-1 α的突变体进行评估
使用位点特异性诱变和经典遗传分析。
一种新的移码表型抑制模型的关键特征
将在提案中测试突变。 第一,需要一个
tRNA和mRNA之间的四核苷酸相互作用将通过以下方法进行测试:
编码移码抑制子基因的位点特异性诱变
tRNA。 第二,该提案将测试预测,
反密码子环稳定近同源解码,
同源解码 第三,它将检验扩张是否只会刺激
由可以在mRNA上滑动的tRNA引起的移码。 最后将
测试是否有相同的机制控制-1移码的抑制
突变。
英文摘要
This proposal concerns the mechanism of +1 translational frameshifting,
both events which occur at special mRNA sites which promote a shift in
reading frame (programmed frameshifting) and phenotypic suppression of
frameshift mutations caused by mutant tRNAs (frameshift suppression).
It has long been thought that these two events were unrelated
mechanistically. The classical model for frameshift suppression
proposes that a mutant tRNA with an expanded anticodon loop reads an
expanded four nucleotide anticodon, causing a shift in the downstream
or +1 direction. Programmed frameshifts by contrast now appear to occur
because of near-cognate decoding of the last in-frame codon, which leads
to a translational error at the next codon shifting reading. The
classical model of suppression now appears invalid, and suppression may
occur by a mechanism similar to programmed frameshifts. This proposal
seeks to further explore the mechanism of programmed frameshifting and
to test whether the near-cognate decoding model for frameshift
suppression is correct.
Yeast molecular genetic analysis will be used to test further the
validity of the near-cognate decoding model for programmed
frameshifting. PCR-based gene disruption will eliminate each of the
low-copy tRNA genes in the yeast genome; in such strains the cognate
codons must be read by a near-cognate isoacceptor, which may induce high
efficiency frameshifting. In some cases frameshifting occurs because
the normal cognate tRNA decodes doo inefficiently; site-specific
mutagenesis will identify the sequences in the tRNAs contributing to
their inefficiency. An in vitro translation system which promotes +1
programmed frameshifting will be used to identify which near-cognate
tRNAs are actually responsible for promoting the shift in frame.
Finally, the basis of frameshift stimulation by a cis-acting mRNA
sequence, and by mutants of elongation factor-1alpha will be assessed
using site-specific mutagenesis and classical genetic analysis.
Key features of a new model for phenotypic suppression of frameshift
mutations will be tested in the proposal. First, the necessity for a
four nucleotide interaction between the tRNA and mRNA will be tested by
site-specific mutagenesis of a gene encodig a frameshift suppressor
tRNA. Second, the proposal will test the prediction that expansion of
the anticodon loop stabilizes near-cognate decoding, and destabilizes
cognate decoding. Third, it will test whether expansion only stimulates
frameshifting by tRNAs which can slip on the mRNA. Finally, it will
test if the same mechanism governs suppression of -1 frameshift
mutations.
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会议论文
TRNA MODIFICATION AND PROGRAMMED TRANSLATIONAL FRAMESHIF
-
批准号:2292682
-
项目类别:
-
资助金额:$4.61万
-
财政年份:1997
-
负责人:Philip James Farabaugh
-
依托单位:
MOLECULAR ANALYSIS OF SITE-SPECIFIC TRANSLATIONAL FRAMES
-
批准号:3277094
-
项目类别:
-
资助金额:$13.5万
-
财政年份:1989
-
负责人:Philip James Farabaugh
-
依托单位:
Molecular genetics of translational accuracy
-
批准号:7896489
-
项目类别:
-
资助金额:$32.76万
-
财政年份:1989
-
负责人:Philip James Farabaugh
-
依托单位:
MOLECULAR BASIS OF TRANSLATIONAL RECODING IN YEAST
-
批准号:2175528
-
项目类别:
-
资助金额:$19.68万
-
财政年份:1989
-
负责人:Philip James Farabaugh
-
依托单位:
The genetics of ribosomal reading frame maintenance
-
批准号:6625800
-
项目类别:
-
资助金额:$32.31万
-
财政年份:1989
-
负责人:Philip James Farabaugh
-
依托单位:
MOLECULAR BASIS OF TRANSLATIONAL RECODING IN YEAST
-
批准号:2175529
-
项目类别:
-
资助金额:$23.17万
-
财政年份:1989
-
负责人:Philip James Farabaugh
-
依托单位:
MOLECULAR BASIS OF TRANSLATIONAL RECODING IN YEAST
-
批准号:2175530
-
项目类别:
-
资助金额:$24.03万
-
财政年份:1989
-
负责人:Philip James Farabaugh
-
依托单位:
MOLECULAR BASIS OF TRANSLATIONAL RECODING IN YEAST
-
批准号:2391900
-
项目类别:
-
资助金额:$24.97万
-
财政年份:1989
-
负责人:Philip James Farabaugh
-
依托单位:
MOLECULAR BASIS OF TRANSLATIONAL RECODING IN YEAST
-
批准号:6179471
-
项目类别:
-
资助金额:$30.81万
-
财政年份:1989
-
负责人:Philip James Farabaugh
-
依托单位:
Molecular genetics of translational accuracy
-
批准号:7417655
-
项目类别:
-
资助金额:$32.3万
-
财政年份:1989
-
负责人:Philip James Farabaugh
-
依托单位:
The genetics of ribosomal reading frame maintenance
-
批准号:7248501
-
项目类别:
-
资助金额:$11.27万
-
财政年份:1989
-
负责人:Philip James Farabaugh
-
依托单位:
Molecular genetics of translational accuracy
-
批准号:7585615
-
项目类别:
-
资助金额:$34.0万
-
财政年份:1989
-
负责人:Philip James Farabaugh
-
依托单位:
SITE-SPECIFIC TRANSLATIONAL FRAMESHIFTING
-
批准号:3277093
-
项目类别:
-
资助金额:$13.16万
-
财政年份:1989
-
负责人:Philip James Farabaugh
-
依托单位:
MOLECULAR BASIS OF TRANSLATIONAL RECODING IN YEAST
-
批准号:6385419
-
项目类别:
-
资助金额:$31.61万
-
财政年份:1989
-
负责人:Philip James Farabaugh
-
依托单位:
MOLECULAR BASIS OF TRANSLATIONAL RECODING IN YEAST
-
批准号:2630925
-
项目类别:
-
资助金额:$26.33万
-
财政年份:1989
-
负责人:Philip James Farabaugh
-
依托单位:
MOLECULAR ANALYSIS OF SITE-SPECIFIC TRANSLATIONAL FRAMES
-
批准号:3277095
-
项目类别:
-
资助金额:$14.4万
-
财政年份:1989
-
负责人:Philip James Farabaugh
-
依托单位:
MOLECULAR ANALYSIS OF SITE-SPECIFIC TRANSLATIONAL FRAMES
-
批准号:3277096
-
项目类别:
-
资助金额:$14.69万
-
财政年份:1989
-
负责人:Philip James Farabaugh
-
依托单位:
The genetics of ribosomal reading frame maintenance
-
批准号:6479044
-
项目类别:
-
资助金额:$32.36万
-
财政年份:1989
-
负责人:Philip James Farabaugh
-
依托单位:
The genetics of ribosomal reading frame maintenance
-
批准号:6868887
-
项目类别:
-
资助金额:$32.31万
-
财政年份:1989
-
负责人:Philip James Farabaugh
-
依托单位:
MOLECULAR ANALYSIS OF SITE-SPECIFIC TRANSLATIONAL FRAMES
-
批准号:3277090
-
项目类别:
-
资助金额:$1.0万
-
财政年份:1989
-
负责人:Philip James Farabaugh
-
依托单位:
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