CiBER-seq dissects genetic networks
Cells integrate environmental signals and internal states to dynamically control gene expression. Muller et al. developed a technique to dissect this cellular logic by linking targeted, genome-wide genetic perturbations with a deep-sequencing readout that quantitatively measured the expression phenotype induced by each perturbation. The method, dubbed CiBER-seq, was able to recapitulate known regulatory pathways linking protein synthesis with nutrient availability in budding yeast cells. Unexpectedly, the authors found that the cellular logic also appears to consider protein production machinery in this decision. By uncovering additional facets of this deeply conserved pathway, the findings demonstrate the utility of comprehensive and quantitative CiBER-seq profiling in mapping the gene networks underlying cellular decisions.
Science, this issue p. eabb9662
Systematically profiling the effects of genetic perturbations is a powerful approach that has revealed the molecular basis for a wide range of biological phenomena. The simple, programmable DNA recognition of CRISPR-Cas9 enables genome-wide genetic analysis in human cells and many other systems. Cas9 is guided by a short RNA to a complementary sequence in the genome, where it can introduce mutations or alter gene expression. Pooled libraries of guide RNAs (gRNAs) that individually target each gene in the genome allow us to introduce genetic perturbations systematically into a population of cells. A key challenge is measuring the phenotypic effects caused by individual guides in these pooled libraries and linking these phenotypes back to the associated gRNA, thereby finding the gene that is responsible.
Molecular phenotypes such as gene expression changes provide a clear and sensitive measure for many cellular processes. We sought a general approach to profile how the expression of a particular gene of interest changed when other genes were perturbed. We began with a library of gRNAs, each disrupting one gene, and linked these guides with an expression reporter containing a guide-specific nucleotide barcode. gRNAs that alter reporter expression will change the abundance of the expressed RNA barcode specifically associated with that guide. Deep sequencing of these expressed barcodes quantifies each of these guide-specific reporter expression effects individually within a pooled, complex population. We have implemented this strategy by combining CRISPR interference (CRISPRi) with barcoded expression reporter sequencing (CiBER-seq).
We used CiBER-seq to profile the responses of several yeast promoters tied to a range of biological functions. Each promoter yielded a distinct pattern of responses that could be understood in terms of its known function and regulation. For example, we rediscover the control of MET6 expression by regulatory ubiquitylation and connect the bud scar protein Cwp1 to other genes required for budding and cytokinesis. Our analysis of the HIS4 promoter, a well-characterized target of the integrated stress response, yielded a range of genetic perturbations that activate this pathway by causing the accumulation of uncharged transfer RNAs (tRNAs). We also uncovered a notable role for tRNA depletion in this response, as impaired tRNA biogenesis activated HIS4 expression through a distinct pathway. In order to understand this regulation, we carried out genetic interaction analysis and looked for quantitative deviations in CiBER-seq profiles caused by the introduction of a second genetic perturbation. We also developed an indirect CiBER-seq approach to measure translational and posttranslational regulation, which both play roles in the signaling pathways upstream of HIS4.
CiBER-seq produces comprehensive phenotypic profiles that offer insights into gene function and regulation. These high-throughput and quantitative phenotypic measurements are also well suited for the systematic measurement of genetic interactions, which contain rich information about the operation of biological processes. This approach can be applied to study a wide range of transcriptional, translational, and posttranslational regulatory responses, and it has the potential to shed light on many areas of biology.
CRISPR-Cas9 gRNA cassettes are linked with transcriptional reporters containing specific barcodes. The RNA-to-DNA ratio for each barcode, measured by deep sequencing, reveals the reporter expression phenotype induced by each gRNA.
To realize the promise of CRISPR-Cas9based genetics, approaches are needed to quantify a specific, molecular phenotype across genome-wide libraries of genetic perturbations. We addressed this challenge by profiling transcriptional, translational, and posttranslational reporters using CRISPR interference (CRISPRi) with barcoded expression reporter sequencing (CiBER-seq). Our barcoding approach allowed us to connect an entire library of guides to their individual phenotypic consequences using pooled sequencing. CiBER-seq profiling fully recapitulated the integrated stress response (ISR) pathway in yeast. Genetic perturbations causing uncharged transfer RNA (tRNA) accumulation activated ISR reporter transcription. Notably, tRNA insufficiency also activated the reporter, independent of the uncharged tRNA sensor. By uncovering alternate triggers for ISR activation, we illustrate how precise, comprehensive CiBER-seq profiling provides a powerful and broadly applicable tool for dissecting genetic networks.
- Mating Study Unlocks the Genetic Code of Attraction - Neuroscience News - March 18th, 2024
- Mindfulness in your DNA? Capacity to be present is partly a function of genetics, study finds - PsyPost - March 18th, 2024
- Genetic testing scam targets Medicare recipients, including those in Spokane - KXLY Spokane - March 18th, 2024
- Some people are genetically predisposed to gain weight. These 5 tips can help - CNN - March 18th, 2024
- SOPHiA GENETICS and The French Kidney Cancer Research Network (UroCCR) Publish Results from Multiyear ... - PR Newswire - March 18th, 2024
- Tissue samples show the deep genetic and cellular impacts of smoking - Medical Xpress - March 18th, 2024
- The regulatory landscape of chromatin accessibility - Nature.com - March 18th, 2024
- Genome assemblies of 11 bamboo species highlight diversification induced by dynamic subgenome dominance - Nature.com - March 18th, 2024
- Research reveals the genetic code behind non-identical twins - Medical Xpress - March 18th, 2024
- How genetic therapies transformed the lives of sickle cell patients - KPVI News 6 - March 18th, 2024
- The Next Wave of Privacy Litigation: The Illinois Genetic Information Privacy Act - Perkins Coie - March 18th, 2024
- State initiates study on genetic cancers, Lynch Syndrome - The Times of India - March 18th, 2024
- How genetic therapies transformed the lives of sickle cell patients - Citizentribune - March 18th, 2024
- Dr Plichta on Areas of Interest for Genetic Testing Research in Breast Cancer - OncLive - March 18th, 2024
- The Genetic History of the Jews Mosaic - Mosaic - March 18th, 2024
- New insights into genetic mechanisms could improve treatment of liver fibrosis - Medical Xpress - March 18th, 2024
- 5 Questions To Ask About The Ethics Of Genetic Data - Forbes - March 18th, 2024
- Couple raising money to fix sons club feet caused by rare genetic condition - 21 Alive News - February 18th, 2024
- What can bulls tell us about men? Genetic discovery could translate to human fertility research - Medical Xpress - February 18th, 2024
- Legislation seeks to bar life insurance companies from purchasing recreational genetic testing data - Delaware First Media - February 18th, 2024
- Family-based study identifies potential new genetic factors linked to Alzheimer's risk in people with African ancestry - National Institute on Aging - February 18th, 2024
- Genetic determinants of micronucleus formation in vivo - Nature.com - February 18th, 2024
- Near-gapless and haplotype-resolved apple genomes provide insights into the genetic basis of rootstock-induced ... - Nature.com - February 18th, 2024
- Accurate and sensitive mutational signature analysis with MuSiCal - Nature.com - February 18th, 2024
- CRISPR, the Genetic Revolution of the 21st Century | OpenMind - BBVA OpenMind - February 18th, 2024
- Genetic control of thermomorphogenesis in tomato inflorescences - Nature.com - February 18th, 2024
- Natural selection and genetic diversity maintenance in a parasitic wasp during continuous biological control application - Nature.com - February 18th, 2024
- Understanding how natural genetic variation contributes to adaptive responses to low oxygen - News-Medical.Net - February 18th, 2024
- Division of Epidemiology and Genetics - Epidemiology and Genetics - University of Southern California - February 18th, 2024
- Genetic study clarifies the link between birth weight and adult morbidity - Medical Xpress - February 18th, 2024
- AI and genetics underpin project to speed up CVD diagnosis and personalise treatment - Hospital Healthcare - February 18th, 2024
- Study Reveals Complex Interaction Between Genetics and Environment in Brain Functioning - Medriva - February 18th, 2024
- Unraveling the Impact of Genetics and Socioeconomic Status on Behavioral and Psychiatric Traits - Medriva - February 18th, 2024
- Genetic Investigation Reveals CETP Gene Variants Linked to Cardiovascular Resilience in Genome-Wide Association ... - Physician's Weekly - December 30th, 2023
- Raha Kapoor's blue eyes remind fans of her great-grandfather, Raj Kapoor; here's what genetics says - IndiaTimes - December 30th, 2023
- Genetic clue to pericarditis inflammation points to promising new treatments - News-Medical.Net - December 30th, 2023
- Certain genetic predispositions may increase risk of atrial fibrillation among those with high alcohol consumption - 2 Minute Medicine - December 30th, 2023
- Understanding the genetic basis of hereditary diffuse gastric cancer - News-Medical.Net - December 30th, 2023
- Genetic engineering was meant to save chestnut trees. Then there was a mistake. - The Washington Post - December 30th, 2023
- Unraveling the mystery of hereditary diffuse gastric cancer - EurekAlert - December 30th, 2023
- What is Genetics? | AMNH - American Museum of Natural History - December 22nd, 2023
- Who killed Shelley Connolly? Genetic genealogy leads authorities to a suspect and a conviction - Alaska's News Source - December 22nd, 2023
- C.S. Lewis, Atheism, and the Genetic Fallacy - Word on Fire - December 22nd, 2023
- FDA approves first genetic test to assess opioid addiction risk using DNA sample - Fox News - December 22nd, 2023
- Genetic blueprint of wild grapes could help breed better grapes - University of California, Davis - December 22nd, 2023
- Unlocking the genetic secrets of grape seedlessness - Phys.org - December 22nd, 2023
- Unlocking the genetic secrets of drought resilience in Persian walnuts - Phys.org - December 22nd, 2023
- FDA approves genetic test to assess patients risk of developing an opioid addiction - KXAN.com - December 22nd, 2023
- How researchers are CReATiNG synthetic chromosomes faster and cheaper - EurekAlert - December 22nd, 2023
- The promise of genetic therapies in sickle cell disease - The Lancet - December 22nd, 2023
- Assessment of genetic diversity, population structure and wolf-dog hybridisation in the Eastern Romanian Carpathian ... - Nature.com - December 22nd, 2023
- Genetics and Heart Disease: What you need to know - IndiaTimes - December 22nd, 2023
- How to become a geneticist - University of Sydney - December 22nd, 2023
- Study sheds light on the ancestry and genetics of Coast Salish woolly dogs | News | Vancouver Island University ... - Vancouver Island University News - December 22nd, 2023
- Human genetics | Description, Chromosomes, & Inheritance - December 13th, 2023
- BASIC GENETICS INFORMATION - Understanding Genetics - NCBI Bookshelf - December 13th, 2023
- Introduction to Genetics - Open Textbook Library - December 13th, 2023
- Clues to preventing Alzheimer's come from patient who, despite genetics, evaded disease Washington University ... - Washington University School of... - December 13th, 2023
- Environmental stress rather than genetics influenced height differences in early Neolithic people: Study - Phys.org - December 13th, 2023
- What is trisomy 18? Why the fatal genetic disorder is in the news and what it's like to get the diagnosis. - Yahoo Life - December 13th, 2023
- Spanish scientists identify the molecular mechanisms controlling the genes involved in proper formation of the heart ... - EurekAlert - December 13th, 2023
- GENEFIT, the First-Ever Fitness Technology to Integrate Personal Genetics with Wearable Tracker Data, Launches to ... - Fitt Insider - December 13th, 2023
- Genetic 'protection' against depression was no match for pandemic stress, finds study of first-year college students - Medical Xpress - December 13th, 2023
- Origin and evolution of the triploid cultivated banana genome - Nature.com - December 13th, 2023
- CellCharter reveals spatial cell niches associated with tissue remodeling and cell plasticity - Nature.com - December 13th, 2023
- How do we get our eye color? A genetics expert reveals the fascinating truth - Fox News - December 13th, 2023
- WVU Today | WVU scientists spice up genetic research through habanero peppers and AI - WVU Today - December 13th, 2023
- Genetic mutations that promote reproduction tend to shorten human lifespan, study shows - Phys.org - December 13th, 2023
- Stanford Scientists Discover Common Genetic Factor That Fends Off Alzheimer's and Parkinson's - SciTechDaily - December 13th, 2023
- Association of genetic risk and lifestyle with pancreatic cancer and their age dependency: a large prospective cohort ... - BMC Medicine - December 13th, 2023
- New bill would block American genetic data from access by companies tied to foreign adversaries - Homeland Preparedness News - December 13th, 2023
- Electric Eels Shocking Ability To Alter The Genetics Of Nearby Animals - Forbes - December 13th, 2023
- "When them genetics kick in its all over" - NBA fans send in rib-tickling reactions as LeBron James attends Zhuri James' volleyball game -... - October 16th, 2023
- David Liu, chemist: We now have the technology to correct misspellings in our DNA that cause known genetic diseases - EL PAS USA - April 7th, 2023
- World Health Day 2023: Understanding the science of Epi-genetics and how to apply it in our daily lives - Free Press Journal - April 7th, 2023
- Genetics - National Institute of General Medical Sciences (NIGMS) - March 29th, 2023
- GENETICS 101 - Understanding Genetics - NCBI Bookshelf - March 29th, 2023
- People always think Im skinny because of good genetics theyre shocked when they see what I used to lo... - The US Sun - March 29th, 2023
- Forensics expert explains 'genetic genealogy' process believed to be used in Kohberger's arrest - KTVB.com - January 6th, 2023
- Idaho student murders: What is genetic genealogy, a tool reportedly used to help capture the suspect? - FOX 10 News Phoenix - January 6th, 2023