Topic 5 - RNAs Beyond the codon Flashcards
What types of RNA would you isolate if you performed a ChIP-like cross-linking experiment followed by IP using an antibody against eukaryotic RNA polymerase II?
RNA chains from genes that are being actively transcribed by RNA polymerase II, i.e. those that are still bound at the cross-linking stage.
Define argonaute
A family of proteins with RNA-binding ability that interact with miRNAs and other short RNAs such as piRNAs. Many argonaute proteins are components of RISC, where their bound miRNAs are used as guides to drive homology-based binding to mRNAs. Argonaute proteins provide the nuclease activity to cleave target mRNAs.
What is a seed sequence?
The shorter sequence, up to 10 bases in length, within a miRNA. Base complementarities between the seed sequence and target mRNA base sequence is essential for miRNA specificity.
If a target sequence is present on different mRNAs expressed in the same cell, what would you predict would be the effect of expressing a miRNA that targets that sequence?
Binding of the RISC–miRNA complex to all the target sequences is possible; therefore you would predict potential translational repression or degradation of all of the mRNAs carrying the target sequence.
Which factors, related to their base sequence, would you need to consider when trying to predict a potential miRNA–mRNA pairing?
There are often mismatches between bases in miRNAs and their target mRNAs. This can mean that target site prediction based upon the base sequence is not reliable. In addition, as very short stretches of homology can drive annealing (as little as 2 nt), the occurrence of possible matches of miRNA seed sequences is also potentially very high.
How might you obtain data on cell-specific miRNA–mRNA interactions in the laboratory?
The mRNA population could be determined through the sequence analysis of isolated mRNA from a cell. To obtain details of the miRNAs within a cell, it is possible to isolate the cellular pool of small RNAs for sequence analysis, or isolate the RNAs that are associated with proteins such as Dicer or components of RISC using immunoprecipitation (rather like the procedure used for ChIP), followed by sequence analysis of the co-immunoprecipitated RNAs.
If a duplicated gene’s mRNA is regulated through its 3′ UTR by a miRNA, how might this feature allow the duplicated gene to acquire a new function?
It is possible that the 3′ UTR sequence itself could mutate so that homology to the original miRNA seed sequences was lost, leading to loss of miRNA regulation, and therefore altering the level of the encoded protein (by losing the translational repression and/or removing the degradation of the mRNA). It is also possible that target sequence mutation could mean the mutated mRNA acquires a new regulator if the new sequence matches a different pre-existing miRNA.
Recall the meanings of the terms homologues and paralogues when used to describe the evolutionary relationships between genes.
Homologues are defined as genes that are related to each other by descent from a common ancestral gene.
Paralogues are genes that are found within the same species that that have arisen from a common ancestor through a duplication event. Paralogues are, therefore, also homologues of each other (as they all share a common evolutionary ancestor).
Modification of local chromatin components is known to be one way in which a cell regulates the transcriptional activity of a gene. Give an example of one such chromatin modification.
Examples could include modifications to core histone proteins such as histone H3. Modifications are seen to various lysine residues (e.g. K4, K9, K27 and K36) and can be in the form of methylation and acetylation.
What are Piwi-associated RNAs (piRNAs)?
Small (
What are long non-coding RNAs (lncRNAs)?
A class of RNA molecules that are over 200 nucleotides in length that do not encode proteins. lncRNA have diverse functions including serving to recruit regulatory proteins to regions of chromatin and act as binding partners through complementary base pairing to other RNA chains to regulate their availability.
What is the molecular function of argonaute proteins?
Argonaute proteins are RNA nucleases that are found as components of RISC and are responsible for degradation of miRNA–mRNA duplexes.
The genomic elimination that occurs during the formation of the new macronucleus does so in an RNA-dependent manner and also requires the function of a Piwi-related argonaute protein. Based upon what you know about the function of argonaute proteins, what does this suggest about the process?
Argonaute proteins are nucleases that play central roles in RISC- and in Piwi-associated degradation of RNAs in cells. This would suggest that one stage most likely involves RNA–RNA hybridisation and degradation.
What molecular process underlies the ability of RNA to detect non-host nucleic acids?
Base-pairing between the host RNA systems (e.g. piRNA) and target non-host RNAs or DNAs.
What are the general classes of proteins that modify chromatin components to modulate transcription?
Examples include histone acetyl transferases (HATs), histone deacetylases (HDACs) and histone methylases.
