Genome Structure & Non-coding RNA

Ultraconserved nonsense: gene regulation by alternative splicing & RNA surveillance

Steven E. Brenner
University of California, Berkeley
Nonsense-mediated mRNA decay (NMD) is a cellular RNA surveillance system that recognizes transcripts with premature termination codons and degrades them. Using RNA-Seq, we discovered thousands of natural lternative splice forms that appear to be targets for NMD. From these, we have been able to gain insight into the mechanism of NMD. Further, we found that this coupling of alternative splicing and RNA surveillance is used as a means of gene regulation. All conserved members of the human SR family of splice regulators have an “unproductive” alternative mRNA isoform targeted for NMD degradation. Preliminary data suggest that this is used for creating a network of auto- and cross-regulation of splice factors. Strikingly, and each lternative splice is associated with an ultraconserved or highly-conserved region of ~100 or more nucleotides of erfect identity between human and mouse--amongst the most conserved regions in these genomes. Further, we found that the most ancient known alternative splicing event is in this family and creates an alternate transcript to be degraded by NMD. Despite conservation since the pre-Cambrian, when the genes duplicate they change their regulation, so that nearly every human SR gene has its own distinctive sequences for unproductive splicing. As a result, this elaborate mode of gene egulation has ancient origins and can involve exceptionally conserved sequences, yet after gene duplication it evolves swiftly and often.
   
Species-specific alternative splicing leads to unique expression of sno-lncRNAs

Li Yang
Shanghai Institutes for Biological Sciences, Chinese Academy of Science
The advent of high-throughput approaches has revealed that a large portion of the mammalian genome is transcribed into long noncoding RNAs (lncRNAs). While many known lncRNAs are polyadenylated, recent work has revealed that a number of lncRNAs are processed in alternative ways without poly(A) tails, which largely failed to be detected by oligo(dT) purification. We have developed an enrichment strategy to purify non-polyadenylated (poly(A)–/ribo–) RNAs used to high throughput sequencing, this strategy has been successfully applied to identify the rich repertoire of non-polyadenylated RNAs in vivo, including a new class of nuclear-enriched intron-derived long noncoding RNAs that are processed on both ends by the snoRNA machinery (sno-lncRNAs). Importantly, the genomic region encoding one abundant class of sno-lncRNAs (15q11-q13) is specifically deleted in Prader-Willi Syndrome (PWS). Although primary sequence analysis revealed that PWS snoRNAs themselves are conserved from human to mouse, PWS sno-lncRNAs are highly expressed in human, but are undetectable in mouse. Importantly, the absence of PWS region sno-lncRNAs in mouse suggested a possible reason why current mouse models fail to fully recapitulate pathological features of human PWS. We further demonstrated that the formation of sno-lncRNAs is often associated with alternative splicing of exons within their parent genes, and species-specific alternative splicing leads to unique expression pattern of sno-lncRNAs in different animals. This study thus further demonstrates a complex regulatory network of coding and noncoding parts of the mammalian genome.
 
Cryo-EM study of the chromatin fiber reveals a double helix twisted by tetra-nucleosomal units
Feng Song1,2*, Ping Chen1*, Dapeng Sun1,2, Mingzhu Wang1, Liping Dong1,2, Dan Liang1,2, Rui-Ming Xu1, Ping Zhu1+, Guohong Li1+
1 National Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing, 100101, China;
2 University of Chinese Academy of Sciences, Beijing, 100049, China
*These authors contributed equally to this work.
+Correspondence to: Guohong Li, Email: liguohong@ibp.ac.cn; Ping Zhu, Email: zhup@ibp.ac.cn
The hierarchical packaging of eukaryotic chromatin plays a central role in transcriptional regulation and other DNA-related biological processes. Here, we report the 11 A° resolution cryo-electron microscopy (cryo-EM) structures of 30 nm chromatin fibers reconstituted in the presence of linker histone H1 and with different nucleosome repeat lengths (NRLs). The structures show a histone H1-dependent left-handed twist of the repeating tetra-nucleosomal structural units, within which the four nucleosomes zigzag back and forth with a straight linker DNA. The asymmetric binding and location of histone H1 in chromatin play a role in the formation of the 30 nm fiber. Our results provide mechanistic insights into how nucleosomes compact into higher-order chromatin fibers.
    
Three-dimensional (3D) genome structure and function

Yijun Ruan
The Jackson Laboratory for Genomic Medicine
400 Farmington Ave, Farmington, CT 06032, USA
It is already more than 10 years after the completion of Human Genome Project. However, the mechanism about how genomic information guides the gene expression in a particular space and time remains to be elucidated. “Structure determines function” is a consensus that we understand the laws of nature. In biology, this consensus is not only applicable to RNA, protein and other small-scale molecules; the same should also apply to macromolecules such as the entire DNA composition of whole genome. With the progress of developing large-scale functional genomic sequencing technologies, we now have the relevant methodologies such as ChIA-PET and Hi-C to map the 3D genome structure and to relate the topological conformation with relevant epigenomic and transcriptional features. Preliminary results from our and others’ studies have shown the influence of higher-order chromatin folding architecture to gene expression outcomes, which provides the basis for further study of genome structure and function. Therefore, the study of the 3D structure and function of the whole genome has become a new trend of development in genomics. This talk will summarize the current advance in this new field 3D genomics. We anticipate that 3D genomics will provide novel insights for understanding the mechanisms of genome regulatory functions and new opportunities of improving human life qualities.
 
Crowding effect and conformation properties of giant DNA

Valentina V. Vasilevskaya
Nesmeyanov Institute of Organoelement Compounds (INEOS), Russian Academy of Sciences, Vavilova ul. 28 Moscow 119991, Russia
Living cells maintain their lives by using a closed medium that contains a rich variety of biomacromolecules under crowded conditions. This molecular crowding significantly affects the structure and function of biomacromolecules. In particular, it is known that, in diluted solutions of, for example, DNA of the T4 bacteriophage, the gyration radius is on the order of 1000 nm and the volume is on the order of 4 × 109 nm3, whereas the gyration radius inside the head of a bacteriophage is about 50 nm and the occupied volume is 5 × 105 nm3. The living cells contain a huge quantity of biomacromolecules, low-molecular-mass salt ions, and other soluble and insoluble components. The features of influence of different components on the conformation of giant DNA will be discussed in this lecture.
 
Fluctuating genome structure and gene regulation

Masaki Sasai
Nagoya University
Recent experimental studies have revealed astonishing features of the three-dimensional (3D) genome structures in various organisms. In order to elucidate the relationship between genome structure and gene regulation, the quantitative modelling of 3D genome structures should be important. A promising approach is to build coarse-grained models of genomes on the basis of the chromosome conformation capture (i.e., 4C, Hi-C or TCC) techniques. By borrowing theoretical ideas to simulate protein folding and fluctuation, we constructed potentials for simulating chromatin movement and performed molecular dynamics calculations of the structural fluctuation of genome in interphase nucleus of budding yeast. By simulating a mutant strain with this method, in which proteins to anchor telomeres to nuclear envelop are suppressed (Taddei et al., 2009, Genome Research), we show that the spatial distributions of the down- and up-regulated genes show their own characteristic changes from wild type, suggesting the importance of the positional regulation of gene expression in nuclei. We also discuss our recent efforts to build a computational model for analyzing structure and dynamics of human genome. The simulated results show the stable distribution of chromosome territories and their fluctuating boundaries in human lymphoblastoid nucleus.
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