Gene organisation, transcription and regulation Flashcards Preview

MCD - Nucleic Acids and Gene Expression > Gene organisation, transcription and regulation > Flashcards

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Features of DNA

- Deoxyribose and phosphate backbone
- Adenine, guanine, cytosine, thymine bases
- C-H bonds make deoxyribose sugar less reactive than ribose
- Stable in alkaline conditions
- Minor grooves make it harder for enzymes to attack DNA


Features of RNA

- Ribose and phosphate backbone
- Adenine, guanine, cytosine, uracil bases
- C-OH bonds make ribose more reactive than deoxyribose
- Unstable in alkaline conditions
- Has larger grooves so more easily attacked by enzymes


Transcription process

- DNA strand unwinds
- Complementary ribonucleotides placed and joined together by phosphodiester bonds
- RNA chain grows one base at a time in 5' to 3' direction
- RNA chain is completed and removed from DNA strand
- DNA strand rewinds


The major classes of RNA and their respective RNA polymerases

rRNA - ribosomal RNA = RNA polymerase 1
tRNA - transfer RNA = RNA polymerase 2
mRNA - messenger RNA = RNA polymerase 3


Gene promoter definition

The DNA sequence at which RNA polymerase binds


Transcription factor definition

A protein that controls the rate of transcription by suppression or activation


Process of Basal Transcription Complex assembly

- Transcription factor (TF) IID binds to TATA DNA sequence
- TF IIA and TF IIB bind to TF IID
- RNA polymerase 2 with TF IIF attached bind.
- TF IIJ, IIE, IIH bind to RNA polymerase 2
(See diagram in notes)
TF IID, TF IIA and TF IIB, RNA polymerase 2 and TF IIF, TF IIE, TF IIH, TFIIJ.


Role of basal transcription complex (BTC)

Allows RNA polymerase 2 to be phosphorylated and carry out transcription
- with no other TFs present, this complex produces a low (basal) level of transcription


Introns definition

Sequences in the gene which are transcribed but edited out of the final mRNA


Exons definition

Segments of the gene which contain sequences that form part of the final mRNA


Sequence of events in pre-mRNA splicing

- U1 binds to splice donor sequence
- U2, U4, U5, U6 all bind and form the splicing complex (spliceosome) which cleaves splice donor sequence
- An "A" residue in the intron is used as the branch point in an intermediate step.
- The "branch" results from cleavage of the phosphodiester bond at the start of the intron
- End of cleaved intron forms bond with brnachpoint "A"
- End of intron is cleaved and intron removed as a "lariat" structure
- The exposed ends of adjacent exon sequences are joined together (ligated)
(see diagram in notes)


Definition of lariat structure

Cleaved intron


Last two nucleotides of exons



First two nucleotides of introns



Sequence at end of introns

15 pyrimidine nucleotides, any Nucleotide, C, A, G


Sequence of splice donor site

End of exon + start of intron = AGGU


Splice acceptor site

End of intron = Pyr15NCAG


Composition of the added 'cap' to pre-messenger RNA

- Formed by hydrolysis of terminal triphosphate or mRNA to a diphosphate
- Further modified by methylation at N7 position in purine ring to form 7-methylguanylate cap


Function of added 'cap' to pre-messenger RNA and how it can be targeted by viruses

- Acts to protect mRNA at 5' end
- Enhances translation of mRNA
- Viruses such as Polio can interfere with recognition of the cap during translation


Process of addition of poly-A tail to pre-messenger RNA and its composition

- Polyadenylation
- Poly A tail added one base at a time
- Added 11-30 bases downstream of AAUAAA sequence which is found in ALL mRNAs
- Essentially just made up of a long tail of A's (AAAA etc)


Mechanisms of RNA interference

- Production of antisense RNA to block mRNA translation
- Production of siRNA from dsRNA
- RNA silencing


Mechanism of antisense RNA being used to block mRNA translation

- Antisense RNA is the reverse of the mRNA being transcribed
- Binds to pre-transcribed RNA to form dsRNA (double-stranded RNA) which cannot be transcribed


Mechanism of dsRNA being used to produce siRNA

- siRNA inhibits RNA that dsRNA is derived from
- Used as a defence mechanism against viral infection
- Small single-stranded RNAs that anneal to viral RNA and degrade them


Meaning of dsRNA

Double-Stranded RNA


Meaning of siRNA

Small Interfering RNA


Process of RNA silencing

- DICER breaks up dsRNA/shRNA into 21-25bp fragments
- siRNA guides endonuclease activity to remove on of the siRNA strands called the passenger strand.
- Retained strand is antisense to the target strand
- Multiprotein RNA-induced silencing complexes (RISCs) are formed
- RISCs recognises antisense siRNA that has bound to target strand and cleaves target
- Target mRNA now cleaved and unable to be translated


What is DICER

An enzyme that cleaves double-stranded RNA to form small double-stranded interfering RNAs (siRNAs)


Meaning of shRNA

Short Hairpin RNA, designed to be used in RNA interference


What are miRNAs

- MicroRNAs
- Involved in regulating other genes


Typical length of a miRNA in nucleotides

18-25 nucleotides


miRNA gene regulation mechanisms

- Sequences end up as siRNA that end up in the RISC
- Mature miRNAs bind to the 3'-untranslated region of mRNAs and subsequently destabilise them, block their translation or both
- Gene knockdown, decreasing the amount of protein created from a gene