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Correlating molecular behavioral phenotypes in a marmoset model of Huntingtons disease

亨廷顿病狨猴模型中分子行为表型的相关性

基本信息

批准号：
10625374
负责人：
ALI H BRIVANLOU
金额：
$ 55.79万
依托单位：
ROCKEFELLER UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-08-01 至 2026-05-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10625374
关键词：
Address Affect Alleles Amino Acids Animal Model Animals Behavior Behavioral Biology Birth Brain imaging Breeding CRISPR/Cas technology Callithrix Callithrix jacchus jacchus Cell Line Cells Code Cognitive deficits Collaborations Communities Complex Corpus striatum structure DNA Derivation procedure Development Developmental Biology Disease Disease Progression Disease model Embryo Exons Experimental Genetics Fertilization in Vitro Frequencies Functional disorder Genes Genome Germ Cells Germ Lines Grant Guide RNA Heritability Human Huntington Disease Huntington gene Imaging technology In Vitro Knock-in Knock-out Laboratories Mental disorders Modeling Molecular Monitor Morula Mus Mutation Nervous System Neurodegenerative Disorders Neuronal Differentiation Neurons Onset of illness Outcome Performance Peripheral Phenotype Pluripotent Stem Cells Population Pre-Clinical Model Primate Diseases Primates Proteins Protocols documentation Reporter Reporter Genes Reproductive Technology Research Retrieval Role Serial Magnetic Resonance Imaging Social Behavior Stretching Surgical Replantation System Technology Transgenic Mice Transgenic Organisms Trinucleotide Repeats Universities Visuospatial autosome behavioral phenotyping behavioral response blastocyst cognitive process developmental disease differentiation protocol embryonic stem cell homologous recombination insight interest knockout gene molecular phenotype mutant nervous system disorder neural neural circuit neurogenetics neurophysiology nonhuman primate optogenetics polyglutamine neurodegenerative diseases post pregnancy prevent single-cell RNA sequencing social deficits stem cell biology stem cells transcriptomics transmission process

项目摘要

ABSTRACT The common marmoset provides a very relevant primate model for understanding the organization of the human nervous system and the diseases that affect it. Like humans, marmosets also demonstrate cooperative social behavior and have advanced cognitive processes, making them of great interest in the field for modeling developmental and psychiatric diseases and their therapies. They are also ideal for multigenerational genetic experiments as they give birth twice a year and mature faster than most primates. However, while the CRISPR/Cas9 system has been used to knockout genes and create knock-ins of single amino acids in a heritable manner in marmosets, it has been a challenge in the field to create germline transmissible models of gene reporters and trinucleotide repeat genes analogous to their murine counterparts. The very low efficiency of homologous recombination (HR) in primates has precluded knocking-in coding sequences by simply injecting Cas9 protein and a guide RNA into embryos during in vitro fertilization (IVF) as is done for creating knockouts. This limitation has prevented modelling of more genetically complex neurological diseases such as Huntington’s disease (HD) and for creating conditional reporters in marmosets, both of which are mainstays in the mouse neurogenetics field. In addition to low HR frequency, other barriers to creating germline transmission of knock- ins include the absence of a well annotated marmoset genome until recently, lack of protocols for derivation of ground state marmoset pluripotent stem cells (cjPSCs), the low percentage of marmoset pregnancies after embryo reimplantation, and a general deficiency of developmental biology expertise in the marmoset field. We propose to harness our labs’ expertise in developmental biology, IVF technologies, and transgenic stem cell biology to overcome this barrier to widespread use of marmosets. We aim to create transgenic knock-in cjPSCs, convert them into ground-state pluripotent stem cells and then inject them into IVF morula to create a chimeric founder marmoset that carries the modified genome. We then aim to screen the transgenic gametes from the founder marmosets to create the F1 progeny and use them to correlate the molecular-behavioral phenotype of HD. As proof-of-principle, we will focus on three knock-in reporter lines to broadly target excitatory, inhibitory, and peripheral neuronal populations. Together, if successful, our aims will result in creation of the first primate model with neuron-specific reporters, establish the marmoset as a valid model of HD, enable access to single- cell transcriptomic changes at the early stages of HD in a primate disease model, and finally correlate these molecular changes with the behavioral phenotype. These aims will provide fundamental insights into the biology of HD and the role of huntingtin protein in different classes of neurons. The outcome of this project will also influence a better understanding of poly-glutamine neurodegenerative diseases that affect humans. In addition, the transgenic marmosets that we generate will be broadly available to the research community and enable new studies into neural circuits, development, behavior, and a wide range of optogenetic applications.

摘要