BIOLOGY 2B03 Lecture Notes - Lecture 20: Schizosaccharomyces Pombe, Genetic Screen, G2 Phase
3 May 2018
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Department
Course
Professor
Genetic'Screen'
Genetic'screen -an'unbiased'search'for'genes'that'are'involved'in'a'particular'
mechanism'
If'you'are'searching'for'something,'you'mutate'every'single'gene,'and'then'you'
look'at'the'phenotypic'effect'of'the'mutation
•
Temperature-sensitive'mutations'allow'researchers'to'change'the'temperature'
and'turn'on'and'off'protein'functions
•
Ex.'Fission'yeast:'S.'pombe
Mutations'in'cell'division'cycle'mutants'(cdc'mutants)'have'1'of'2'effects'
on'a'cell:'either'cells'stay'in'the'G2'phase'too'long'and'end'up'longer'or'
they'enter'the'mitosis'phase'to'early'and'end'up'too'small'
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From'this'we'can'determine'that'with'more'cdc2'you'get'the'wee'
phenotype'and'without'cdc2'the'cell'fails'to'divide'
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This'suggests'that'Cdc2'is'a'key'regulator'for'entry'into'mitosis'and'
promotes'cell'division
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In'fact'Cdc2'with'Cdc13'forms'a'heterodimer,'therefore'Cdc2'is'the'CDK'
of'the'MPF
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Cdc13:'
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Cdc25:'
Deficit'results'in'elongated'cells''
§
Excess'results'in'small'cells'
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Most'likely'a'activator'of'MPF'and'promotes'entry'into'M-phase'
§
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Wee1:'
Excess'results'in'elongated'cells'
§
Deficit'results'in'small'cells'
§
Most'likely'an'inhibitor'of'MPF'and'delays'entry'into'the'M-phase
§
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Wee1'and'Cdc25'phosphorylate'and'dephosphorylate'tyrosine'15'
inactivating'and'activating'MPF'activity'respectively'
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Functional'complementation'
The'principle' of'this'technique'is'to'screen'through'genes'to'identify'one'that'
can'rescue'the'phenotype'caused'by'a'mutation'and'restore'wild-type'
behavior'
•
To'test'this'genes'are'added'back'at'random'to'restore'the'ability'of'the'cell•
A'cDNA'library -a'collection'to'genes'created'for'an'organism'and'stored'in'
bacterial'cells'
The'library'is'created'by'isolating'mRNAs'and'using'reverse'transcriptase'to'
make'DNA'copies'of'the'messages
•
Ex.'Budding'Yeast:'S.'cerevisiae
The'mutation'causes'the'cell' to'arrest'in'G1,'so'the'cell' forms'the'
daughter'but'fails'to'proceed'into'S-phase
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By'doing'the'addition'of'various'genes,'it'is'determined'that'Cdc28'is'the'
mutated'gene
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By'supplying'the'mutated'budding'yeast'with'cdc2,'the'cell' functions'can'
be'saved
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Therefore,'cyclin-CDK'complexes'are'homologous''
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Module'9:'Lecture'2
Tuesday,' April'3,'2018
12:57'PM
Document Summary
Genetic screen - an unbiased search for genes that are involved in a particular mechanism. If you are searching for something, you mutate every single gene, and then you look at the phenotypic effect of the mutation. Temperature-sensitive mutations allow researchers to change the temperature and turn on and off protein functions. From this we can determine that with more cdc2 you get the wee phenotype and without cdc2 the cell fails to divide. This suggests that cdc2 is a key regulator for entry into mitosis and. This suggests that cdc2 is a key regulator for entry into mitosis and promotes cell division. In fact cdc2 with cdc13 forms a heterodimer, therefore cdc2 is the cdk of the mpf. Most likely a activator of mpf and promotes entry into m-phase. Most likely an inhibitor of mpf and delays entry into the m-phase. Wee1 and cdc25 phosphorylate and dephosphorylate tyrosine 15 inactivating and activating mpf activity respectively.
