Dorris Neuroscience Center
Faculty, Graduate Program
Adjunct Associate Professor, Department of Neurosciences, University of California San Diego
Advancing Stem Cell Technology to Study Genomes and the Brain:
Generating Mice from iPS Cells: During development, undifferentiated stem cells generate the millions of distinct cell types in the body in a choreographed sequence that is unidirectional. Until recently, differentiated mammalian cells were thought to have irreversibly lost the ability to generate less differentiated cell types, for unknown reasons. Thus embryonic stem cells and somatic stem cells were assumed to be unique resources to generate more differentiated cell types in vitro. Reprogramming by transient expression of pluripotency factors can restore differentiated cells to a state of developmental potency that resembles embryonic stem (ES) cells, however, the first generations of induced pluripotent stem (iPS) cells differed from ES cells in that they could not generate live adult mice. We recently developed a reprogramming method that restores differentiated fibroblasts to full pluripotency such that we can produce live mice derived entirely from differentiated cells, without using oocytes. These fully pluripotent iPS cells and iPS mice allow us to examine the functional stability of iPS-derived tissues and to probe the genomic stability of differentiated cells. Applying this technology to human cells should allow us to generate improved iPS cell lines. One goal is to use these cells to generate improved models of disease, in particular, inherited human neurological diseases for which no in vitro models exist.
Control of genome stability during reprogramming and in iPSCs: Reprogramming by the cytoplasm of the egg, or by direct expression of transcription factors requires genome-wide remodeling of DNA. This occurs by mechanisms that are poorly understood. Recent studies in human iPS cells suggested that these cells may necessarily harbor increased numbers of mutations compared to ES cells. We recently performed whole genome sequencing on several mouse iPSC lines that have been shown to be fully reprogrammed because they can generate fertile adult mice. In contrast to the reports of others, we find a very low burden of genomic structural changes in these lines. This shows that reprogramming need not be highly mutagenic and opens the door to identifying methods to produce iPS cell lines with minimal genome instability. In addition, we identified some mutations that arose in the donor cell lineage during somatic differentiation. This result, combined with the low mutation rate we observed will allow us to use reprogramming to identify the source and scope of somatic mutations arising in different tissues and identify optimal tissue sources for human iPS cell lines.
Generating Cell Lines from Neurons: Brains are composed of an extraordinary diversity of neurons, which are primarily generated at birth and are maintained without cell division for the life of an individual. Neurons do not divide and have not been shown to generate tumors, for unknown reasons. The inability to generate cell lines from neurons precludes a number of important studies including analyses of neuronal genomic stability in differentiation and disease, and has impeded the generation of appropriate in vitro models of disease. Previously we were the first to generate cloned mice and ES cell lines from differentiated olfactory sensory neurons using reprogramming by nuclear transfer. A current goal is to generate cell lines from neurons to identify unknown irreversible changes to neuronal DNA and to generate the best possible models of neurological diseases such as autism, schizophrenia, Parkinson’s disease and developmental diseases arising from inherited defects in genome maintenance and epigenetic alterations.
Modeling Neurological Disease Using Reprogramming: Reprogramming may be used to generate neurons in vitro in two ways. First, non-neuronal tissues from mice or humans affected with a particular disease may be reprogrammed to pluripotency and the resulting iPSCs or SCNT-ESCs may be subsequently differentiated into cells of the neural lineage. This approach may be useful for modeling neurodevelopmental diseases – from rare inherited monogenic disorders to more common disorders such as autism or schizophrenia. A second approach is to directly reprogram skin cells or other lineages to neurons without transiting through any precursor states using transcription factors or microRNAs. Our laboratory is exploring the capabilities and limitations of both approaches using mouse and human models.
Generating Neuronal Diversity and Connectivity in the Olfactory System. Exciting recent advances in stem cell technology that suggest it may be possible to generate neurons in vitro to model disease or for use in cell replacement therapy. However, little is known about the extent of genetic diversity of neurons of the same subtype, or how diverse neurons are wired together to form a functional neuronal circuit. We exploit the genetic tractability and anatomic stereotypy of the olfactory system to generate gene targeted mice in which we can label and modify specific neuronal circuits involved in detecting smells. We are currently mapping the links between the nose and cortical brain regions involved in responding to odors. Identifying the patterns of gene expression and synaptic connectivity of defined neuronal populations will be important to assess the potential utility of iPS derived neurons in vitro and in transplantation experiments and should shed light on design principles of functional neuronal circuits.
B.S., Economics, Duke University, 1991
Ph.D., Immunology, Stanford University, 1998
2011 – present Associate Professor, Department of Cell Biology, The Scripps Research Institute, La Jolla, CA.
2010 – present Investigator, Dorris Neuroscience Center, The Scripps Research Institute, La Jolla, CA.
2010 – present Assistant Adjunct Professor, Department of Neurosciences, University of California, San Diego, La Jolla, CA.
2006 – 2011 Assistant Professor. Department of Cell Biology, The Scripps Research Institute, La Jolla, CA. 2006 – present Member, Neurosciences Graduate Program, University of California, San Diego, La Jolla, CA.
2003 – 2005 Associate Research Scientist, Center for Neurobiology and Behavior. Howard Hughes Medical Institute, Columbia University Medical Center, New York, NY. Laboratory of Richard Axel, M.D.
1998 – 2003 Postdoctoral Research Fellow, Center for Neurobiology and Behavior. Howard Hughes Medical Institute, Columbia University Medical Center, New York, NY. Laboratory of Richard Axel, M.D.
1997 – 1998 Postdoctoral Research, Department of Microbiology and Immunology, Howard Hughes Medical Institute, Stanford University, Stanford, CA. Laboratory of Mark M. Davis, Ph.D.
1992 – 1997 HHMI Pre-doctoral Fellow, Department of Microbiology and Immunology, Howard Hughes Medical Institute, Stanford University, Stanford, CA. Laboratory of Mark M. Davis, Ph.D.
1990 – 1991 Research Assistant, National Institute of Environmental Health Sciences. National Institute of Health. Research Triangle Park, NC. Laboratory of Michael A. Resnick, Ph.D.
2011 – 2012 Donald E. and Delia B. Baxter Foundation Faculty Scholar
2007 – 2011 Pew Scholar in the Biological Sciences
Boland MJ, Hazen JL, Nazor KL, Rodriguez AR, Gifford W, Martin G, Kupriyanov S, Baldwin KK. Generation of mice derived from induced pluripotent stem cells. Journal of Visualized Experiments (manuscript accepted, 2012).
*Quinlan, AR, *Boland, MJ, Leibowitz, ML, Shumilina, S., Pehrson, SM, *Baldwin, KK, and *Hall, IM. Genome sequencing of mouse induced pluripotent stem cells reveals retroelement stability and infrequent DNA rearrangement during reprogramming. Cell Stem Cell. 2011 Oct 4;9(4):366-73. these authors contributed equally.
Ghosh, S., Larson, S., Hefzi, H., Marnoy, Z., Cutforth, T., Dokka, K., and Baldwin, KK. Sensory maps in the olfactory cortex defined by long-range viral tracing of single neurons. Nature. 14 April 2011; 472(7342):217-20.
*Boland MJ, *Hazen JL, *Nazor K, Rodriguez AR, Gifford W, Martin G, Kupriyanov S, and Baldwin KK. Adult mice generated from induced pluripotent stem cells. Nature. 03 September 2009. 46:91-96. * these authors contributed equally.
*Eggan, K., *Baldwin, KK., Tackett, M., Osborne , J., Gogos, J., Chess, A., Axel, R. and Jaenisch, R. Mice cloned from olfactory sensory neurons. Nature. 04 March 2004. 428: 44-49. * these authors contributed equally.
*Tasic, B., *Nabholz, C.E., Baldwin, KK., Kim, Y., Rueckert, E.H., Ribich, S.A., Cramer, P., Wu, Q., Axel, R. and Maniatis, T. Promoter choice determines splice site selection in protocadherin alpha and gamma pre-mRNA splicing. Molecular Cell. 2002. 10(1): 21-33. * these authors contributed equally.