Lecture 8 Flashcards
What are the functions of RNAs?
mRNA - messenger rna with 5’ cap, introns removed by rna splicing and a poly(A) tail
pre-mRNA - an mRNA precursor containing introns and not cleaved at poly(A) site
hnRNA - heterogenous nuclear RNA, include pre-mRNAs and RNA processing intermediates
snRNA - five small nuclear RNAs that function in the removal of introns from pre-mRNA
pre-tRNA - tRNA precursor containing additional transcribed bases at the 5’ and 3’ ends, some contain intron in anticodon cap
pre-rRNA - precursor to mature rRNAs, mature rRnas are processed from this long molecule by cleavage
snoRNA - small nucleolar rnas, they base pair with complementary regions of pre-mRNA molecules, directing cleavage of the RNA chain and mod of bases during maturation of rRNAs
siRNA - short interfering RNAs, cause cleavage of the target RNA, leading to degradation
miRNA - microRNAs, base pair with mRNAs, inhibits translation of the target mRNA and targets it for degradation
What is the basic process of turning pre-mRNA into mRNA?
pre-mRNA is capped, then spliced then polyadenylated before being exported to the cytoplasm:
- splicing: a large ribonucleoprotein spliceosome complex catalyzes the joining of two exons and the removal of introns
- network of interactions between SR proteins, snRNPs, and
splicing factors forms a cross-exon recognition complex that
specifies correct splice sites
What does alternative splicing of mRNA yield?
significant variety of different mRNAs from the same gene in different cell types or at different developmental stages, some of the proteins generated by alternative splicing have drastically different activities
Function of alternative splicing in the perception of sounds of different frequencies system?
*(a) Chicken cochlea:
*5-mm long tube containing an epithelium of auditory hair cells that are tuned to a gradient of vibrational frequencies from 50 Hz at the apical end (left) to 5000 Hz at the basal end (right)
*Individual “hair cell” neurons – respond most strongly to a specific sound frequency by expressing a mixture of specific Ca2+-activated K+ channel isoforms
*(b) Ca2+-activated K+ channel:
*Seven transmembrane α helices (S0–S6) form the K+ channel.
*Cytosolic domain –
*includes four hydrophobic regions (S7–S10).
*regulates opening of the channel in response to intracellular Ca2+ concentrations.
*Ca2+ concentration at which the channel opens determines the frequency of membrane potential oscillation- frequency to which the cell is tuned.
*Channel isoforms –
*encoded by alternatively spliced mRNAs produced from the same primary transcript
*alternative exons used at eight regions in the mRNA – 576 possible isoforms [Red numbers – regions where alternative splicing produces different isoforms]
*respond to different frequencies by opening at different Ca2+ concentrations
How many possible isoforms of a membran Ca2+ activated K+ channel from a single mRNA?
576
Alternative isoforms of the drosophilia Dscam gene system?
*Dscam isoform expression in Drosophila neurons:
*Most extreme example of regulated alternative RNA processing yet discovered
*Helps to specify tens of millions of different specific synaptic connections between neurons in the Drosophila brain
*(bottom) Dscam neuron cell-surface protein:
*10 different immunoglobulin (Ig) domains (ovals)
*6 different fibronectin type III domains (rectangles)
*One transmembrane domain (yellow)
*C-terminal cytoplasmic domain
*Fully processed mRNA contains:
*One of 12 Ig2 exons
*One of 48 Ig3 exons
*One of 33 Ig7 exons
*One of 2 transmembrane exons
*Alternative splicing – can generate 12 x 48 x 33 x 2 = 38,016 possible isoforms
What are some splicing changes observed in people living with autism spectrum disorder? (ASD)
*RNA-seq revealed that cassette exons and microexons (3–27 nucleotides) are much more common forms of alternative RNA splicing in the normal human central nervous system than alternative 5′ and 3′ splice sites, retained introns, or complex events.
*In one-third of patients with autism spectrum disorder (ASD), more skipping of cassette exons and microexons are observed than in normals.
*Abnormalities in the frequency of splicing of alternative 5′ and 3′ splice sites are observed only rarely in ASD.
Complex splicing events, while present, occur at too low an incidence to determine if their incidence is altered significantly in ASD patient.
What is sxl?
RNA binding protein, acts as a suppressor of splicing
What is tra?
RNA binding protein, acts as an activator of splicing
What is dsx?
transcription activator/repressor
What regulates alternative splicing in drosophila?
proteins
Where does splicing occur in sex determination in drosophila?
exons and introns, divided by female and males
How does sxl protein play a role in sex determination in drosphila?
Sex-lethal protein: intronic splicing silencer - present only in female embryos (no function sxl protein in males), it binds to a pre-mRNA sequences near the 3’ end of the intron betweene xon 2 and exon 3, sxl blocks U2AF and U2 snRNP association with 3’ splice site in males
female: deletes exon 3 which contains premature stop codon and keeps exon 2/4
male: keeps exon 3, which contains the premature stop codon
How does tra play a role in sex determination in drosophila?
sxl blocks tra exon 1-2 splicing
only female embryos produce function tra protein
tra promotes specific dsx splicing
What role does Dsx play in sex determination in drosophila?
females: cooperative binding of tra protein and rbp1 and tra2 sr proteins activates exon 2-4 splicing and cleavage (polyadenylation) at the 3’ end of exon 4, expresses female dsx isoform
males: embryos lack functional tra, sr proteins do not bind to exon 4, exon 3 is spliced to exon 5, expresses male dsx isoform
