Основы биологии_Лекция-11 (2013)
.pdfФБМФ МФТИ Основы биологии
Лекция 11
2013
Транскрипция. Особенности генома эукариот.
• РНК-полимеразы, механизмы транскрипции у прокариот и эукариот.
•Регуляция активности промоторов и терминаторов. Аттенюация.
•Процессинг у прокариот и у эукариот, альтернативный
сплайсинг, транссплайсинг.
•Строение эукариотического генома. Повторы, сателлитные ДНК. Уникальные последовательности
генома.
•Подвижные элементы генома. Прерванные гены
эукариот.
1
Контроль реализации генетической информации на уровне транскрипции
DNA |
Environmental change |
|
|
RNA |
Turn gene(s) on/off |
|
protein
Proteins to deal with new environment
Very important to:
1.express genes when needed
2.repress genes when not needed
3.Conserve energy resources; avoid expressing unnecessary/detrimental genes
Транскрипция у прокариот
•Operons
Groups of related genes transcribed by the same promoter
•Polycistronic RNA
•Multiple genes transcribed as ONE TRANSCRIPT
•No nucleus, so transcription and translation can occur simultaneously
2
Синтез РНК РНК-полимеразой•
•
•RNAP binds, melts DNA
•Nucleosides added 5’ 3’
Типы РНК
•Messenger RNA (mRNA) – genes that encode proteins
•Ribosomal RNA (rRNA) – form the core of ribosomes
•Transfer RNA (tRNA) – adaptors that link amino acids to mRNA during translation
•Small regulatory RNA – also called noncoding RNA
3
Структура бактериальной РНК-полимеразы
•RNA polymerase
–4 core subunits
–Sigma factor (σ)– determines promoter specificity
–Core + σ = holoenzyme
–Binds promoter sequence
–Catalyzes “open complex” and transcription of DNA to RNA
Структура промотора
• Sigma factors recognize consensus -10 and -35 sequences
4
Консервативные последовательности промоторов
TTGACA |
TATAAT |
Deviation from consensus -10 , -35 sequence leads to weaker gene expression
Варианты сигма-факторов
Sigma subunit |
Type of gene controlled |
# of genes controlled |
|
|
|
|
|
70 |
RpoD |
Growth/housekeeping |
~1000 |
54 |
RpoN |
N2; stress response |
~15 |
S |
RpoS |
Stationary phase, virulence |
~100 |
S |
RpoH |
Heat shock |
~40 |
F |
RpoF |
Flagella-chemotaxis |
~40 |
32 |
RpoE |
Extreme heat shock, unfolded proteins |
~5 |
|
? |
||
|
FecI |
Ferric citrate transport |
~5 |
E. coli can choose between 7 sigma factors and about 350 transcription factors to fine tune its transcriptional output
An Rev Micro Vol. 57: 441-466 T. M. Gruber
5
Ключевые цифры
•In log-phase E. coli:
–~4000 genes
–~2000 core RNA polymerase molecules
–~2/3 (1300) are active at a time
–~1/3 (650) can bind σ subunits.
•Competition of σ for core determines much of a cell’s protein content.
Терминация транскрипции
•Bacteria need to end transcription at the end of the gene
•2 principle mechanisms of termination in bacteria:
–Rho-independent (more common)
–Rho-dependent
6
Rho-независимая терминация
• Termination sequence has 2 features: Series of U residues
GC-rich self-complimenting region
•GC-rich sequences bind forming stem-loop
•Stem-loop causes RNAP to pause
•U residues unstable, permit release of RNA chain
Rho-зависимая терминация
•Rho is hexameric protein
•70-80 base segment of RNA wraps around
•Rho has ATPase activity, moves along RNA until site of RNAP, unwinds DNA/RNA hybrid
•Termination seems to depend on Rho’s ability to “catch up” to RNAP
•No obvious sequence similarities, relatively rare
7
Аттенюация
•Attenuator site = DNA sequence where RNAP chooses between continuing transcription and termination
•trp operon
–4 RNA regions
for basepairing
–2 pairs w/ 1 or 3
–3 pairs w/ 2 or 4
–Concentration of Trp-tRNATrp determines fate of attenuation
–At high Trp conc, transcription stops via Rho-independent
Транскрипция у эукариот
8
Orphanides, Cell 2002
•Контроль за реализацией генетической информации у эукариот идёт на всех этапах
Activation of |
Initiation of |
gene structure |
transcription |
9
Структура эукариотического гена
•“The entire nucleic acid sequence that is necessary for the synthesis of a functional polypeptide or RNA molecule”
Основные элементы транскрипции
•Initiation, elongation, termination
•Catalyzed by RNA polymerase
–“Transcription bubble”: DNA transiently separated into single strands
–One strand is used as a template
–Unwinding point & rewinding point
–Rate 40 nucleotides/second at 37 for bacteria
•RNA polymerase
–Many subunits: catalytic site, CTD with (YSPTSPS)n
–pol I, pol II, pol III
10