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CMB2001 EXAM WITH VERIFIED ANSWERS AND QUESTIONS
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CMB2001 EXAM WITH VERIFIED ANSWERS ANDQUESTIONS
Describe how H3Lys9me heterochromatin is formed - ANSWER H3Lys9me
heterochromatin formation: Lys9 deacetylated by HDAC. Suvar39 then methylates
lys9. Methylated lys9 is recognised by HP1 and then HP1 binds. HP1 has
chromodomain - chromodomains often recognise and bind to methylated lys
residues. The chromodomain on HP1 is specific for H3Lys9me (with di or
tri-methylated lys 9). The binding of HP1 is thought to compact nucleosome arrays
as HP1s join together.
What is gene expression? - ANSWER Process by which DNA is decoded to
proteins
Causes protein synthesis (DNA to RNA by transcription, RNA to protein by
translation)
It is highly regulated
What is transcription? - ANSWER ds DNA to ss mRNA
Template strand copied 5' to 3'
Describe transcription in prokaryotes - ANSWER RNA polymerase with sigma
factor 70 (holoenzyme) makes contact with -35 and -10 promoter region (upstream
from transcription start site). This forms a closed complex.
The DNA is opened (open complex) over the transcription start site and RNA pol
uses a template strand to begin making a copy (initiation). Sigma factor 70 is
released and the mRNA strand is elongated 5' to 3'.
What does a promoter do? - ANSWER Initiate and control transcription when RNA
pol binds
What are +1, -10 and -35 sequences on prokaryote promoter? - ANSWER + 1 is the
first base that is used as a template
-10 and -35 are consensus sequences which are the strongest promoters and are
,upstream. Both hexamers
Sequences are very defined in prokaryotes
Describe eukaryotic promoters (go into detail on core and regulatory regions) -
ANSWER There is a core (basal region) around transcription start site, and a
regulatory region upstream.
TATA box at -30 and initiator at +1 are found on the core region. There is also
DPE at +30 and BRE at -35. Often core regions will not have all of these elements
The regulatory region has proximal elements (UAS and URS) and distal (enhancer
and silencer). UAS and enhancer and activator binding sites which increase
transcription. URS and silencer are repressor binding sites which decrease
transcription.
Sequences are less defined than they are in prokaryotes
What are CpG islands? - ANSWER Regions with high frequency of CG sequences
(CpG islands). These CG sequences have C residues that are hypomethylated
which causes decreased transcription (due to less initiation)
What do reporter genes do? - ANSWER Encode proteins whose levels can be
easily measured eg GFP and LacZ
The amount of reporter protein is proportional to gene expression (activity)
Reporter genes can be added to plasmids which cause recipient cells to have
reporter gene expression
Contrast types of RNA pol in prokaryotes and eukaryotes - ANSWER In
prokaryotes there's only one
In eukaryotes there are at least 3 main ones
Similar structure but bacterial has 2 alpha, 1 beta, 1 beta prime and one omega
subunit. Yeast RNA pol II has 12 subunits.
What are types of RNA pol in eukaryotes? - ANSWER RNA pol I - transcribe
rRNA (28S, 18S and 5.8S). Found in nucleolus
RNA pol II - transcribes mRNA (and snRNA and miRNA). Found in nucleus
,RNA pol III - transcribes tRNA (and 5S, U6 and 7S RNA). Found in nucleus
RNA POL II IS THE MAIN ONE
What are general transcription factors (GTFs)? - ANSWER TFII A,B,D, E,F and H
All but TFIIB are made of complexes made of multiple polypeptides.
They are all specific to RNA pol II and form a complex on TATA box and recruit
RNA pol II to promoter and direct initiation and start site
Used at every promoter
Explain how PIC (Pre initiation complex) is assembled - ANSWER TFIID binds to
TATA box, then TFIIA and subsequently TFIIB (TFIIB is recruited by D) join.
TFIIB acts as a signal to allow RNA pol II to bind. RNA pol II binds and is
associated with TFIIF which therefore also binds. TFIIE and TFIIH then join.
TFIIH has helicase activity that is powered by ATP hydrolysis. This separates the
template strand at the start site which forms an open complex.
RNA pol II can now start transcription. As this begins RNA pol II is
phosphorylated (a lot) on the C terminal domain (CTD) (which is found on the end
of the beta prime subunit)
TFIID and A may stay behind. B, E and H are released. F moves with RNA pol II
along the template
Describe TFIIH - ANSWER Has 9-10 subunits
Has 2 parts: CORE and CAK
CAK contains a kinase that phosphorylates CTD on RNA pol II.
Also contains ATPase called XPB which causes promoter melting
Describe TFIID and its components - ANSWER Made of TATA binding protein
(TBP) and TBP associated factors (TAFs).
TBP is in the centre. It can cause assembly of PIC kn promoter with TATA box but
not without
TAFs promote interactions of TFIID with basal promoter where there is no TATA
, box (TAF 1 and 2 at initiator, TAF 6 at DPE). Allows TFIID to interacts where
there are no TATA box
It is very well conserved between organisms and has a trilobular structure
Which side is upstream, and which side is downstream? - ANSWER Upstream
means to the left of in diagram, and downstream means to the right
What happens if there is basal promoter w/ or w/ out UAS (upstream activating
sequence)? - ANSWER w/: low/no txn
w/ out: activation therefore high txn
What are common sequence elements and what do they do? - ANSWER GC box,
octamer, and CAAT box.
These are located near core promoter and bind activators that are abundant and
constantly active
What are response elements and what do they do? - ANSWER SRE and HSE -
these bind activators that have controlled / induced activity by specific stimuli. SRE
binds serum response factor and is induced by growth factors, HSE binds heat
shock factor and is induced by heat shock
What is combination control? - ANSWER Where the type and combination of
enhancer/UAS elements (these are often sequence and response elements) induce
different types of txn (eg HSE element may be present during heat shock which
would cause more txn than promoter w/ out HSE)
Describe enhancers - ANSWER Enhancers can work from any location/orientation
from the promoter region. Models either say that enhancer moves along DNA to
RNA pol (tracking), or DNA loops which brings enhancer to RNA pol. Looping is
more likely (almost sure that tracking does not occur)
Describe eukaryotic activators - ANSWER Are modular, meaning they are made of
separable activation and DNA binding domains - these are joined by flexible
protein domains. Eg, activation domain for GAL4 can be joined to separate DNA
binding domain from different protein and still function as an (possibly different)
activator
What is GAL4? - ANSWER Transcriptional activator protein in yeast. It regulates
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