lncRNA Xiaole Shirley Liu STAT115, STAT215, BIO298, BIST520
Outline • Last lecture, short non-coding RNAs: miRNA, siRNA, piRNA • Today: long non-coding RNAs (lncRNAs) • Systematically discovery of lncRNA • Function of lncRNA
ChIP-seq • lncRNA are mostly polII transcribed, and has similar histone mark profiles as protein-coding genes. • Guttman et al, Nat 2009 use active promoter and Pol2 elongation mark to find lncRNAs, know the TSS and direction of transcription
RNA-seq • Mostly not investigated on microarrays before • Many LncRNAs are spliced and have intervening expression pattern with coding genes
LncRNA Functions • Similar to coding RNAs, lncRNA is transcribed by pol2, regulated by TFs, and have polyA tails • Expression levels of lncRNA is often lower than coding RNAs • Gradually finding very important functions of some lncRNAs, some overlapping functions
lncRNA Signaling • Xist and X-inactivation • In vertebrate genomes, male has XY, female has XX but only one of the X is active
lncRNA as miRNA Decoy • Poliseno et al, Nat 2011
LincRNA as Scaffold for Transcription Repression • HOTAIR and PRC2 Tsai et al, Science 2010
Map lncRNAs Associated with Chromatin Factors • Many chromatin factors have RNA-binding domains, functions modulated by lncRNA • RIP-seq of chromatin factors • Downstream analysis?
Map lncRNA Occupancy • ChIRP: Chromatin Isolation by RNA Purification (ChIRP) • Chu et al, Mol Cell 2011
Upstream Antisense RNA • Pervasive upstream antisense RNA Core et al, Science 2008
Enhancer RNA • Weak level of Pol II and RNA transcripts observed at distal TF binding sites Kim et al, Nat 2010
LincRNA as Enhancers • Some LincRNAs promote nearby gene trx Orom et al, Cell 2010
Potential Functions of lncRNAs
Summary • Pervasive ncRNA transcripts exist in the genome, carrying diverse functions from epigenetic, co-transcriptional, and post-transcriptional regulatory roles • NGS enables the discovery and better understanding of these ncRNA and their mechanisms
Grant Proposal: Specific Aims • Grant title and your name • Introductory paragraph (~1/4 page) • A is very important in bio/medicine/disease • Recent development in A has made some really significant findings or improvements • However, something is still lacking or not known about A • The central hypothesis of this proposal is … • Therefore, we plan to do investigate / develop … for … goals
How to Write a Specific Aims Page • Specific Aims (1/3 to ½ page) • To achieve the goals, we plan to: • Aim 1: profile the genome-wide xxx of xxx • 1.1: establish xxx • 1.2 • Aim 2: develop a computer algorithm or knowledge base • 2.1: model xxx • … • Aim 3: identify the mechanism of xxx • …
Specific Aims • Sound like you can definitely do it • Do not use words that sounds like a fishing expedition such as try, explore (find) or words that sound too trivial such as download (integrate) • Try not to let your aims depend on each other • Propose as many aims as years for the grant
How to Write a Specific Aims Page • Last paragraph (< ¼ page) • What’s novel about our approach • Deliverables (what will the scientific community see at the end) • A software, database, map, mechanism, resource • What’s the (potential) significance about our proposal • Best possible outcome
