Zhaolan (Joe) Zhou, Ph.D.

1. Rett syndrome is a debilitating neurodevelopmental disorder caused by mutations in the gene encoding methyl-CpG binding protein 2 (MeCP2). One characteristic feature of Rett syndrome is the regression of learned motor and language skills after 6-18 months of normal development. The onset of the disease coincides with synaptic maturation driven by sensory experience in humans. As a postdoctoral fellow, I started to pursue the role of MeCP2 in neuronal activity-dependent gene regulation and investigated the possibility that defective experience-dependent synaptic maturation underlies the pathogenesis of Rett syndrome. I found that MeCP2 is selectively phosphorylated in the brain in a neuronal activity-dependent manner and this event mediates dendritic morphogenesis and spine maturation. Sponsored by an NIH Pathway to Independence Award (K99/R00), I went on and led a research team finding that MeCP2 regulates gene expression programs and neuronal development in a brain region, neuronal cell type, and age-specific manner. My lab also found MeCP2 plays a necessary and sufficient role in GABAergic interneurons to maintain neural network integrity, providing the cellular origin driving auditory event related potentials (ERPs) and paving the way to investigate Rett syndrome pathogenesis in defined cell types of interest.

a. Zhou Z, Hong E, Cohen S, Zhao W, Ho SY, Chen W, Savner E, Hu L, Steen J, Weitz C and Greenberg ME* (2006). Brain-specific phosphorylation of MeCP2 regulates activity-dependent Bdnf transcription, dendritic growth, and spine maturation. Neuron, 52: 255–269. PMCID: PMC3962021.

b. Wang IT, Reyes AR, and Zhou Z* (2013). Neuronal morphology in MeCP2 mouse models is intrinsically variable and depends on age, cell type, and Mecp2 mutation. Neurobiology of Disease, 58C: 3-12. PMCID: PMC3748238.

c. Zhao YT, Goffin D, Johnson BS and Zhou Z* (2013). Loss of MeCP2 function is associated with distinct gene expression changes in the striatum. Neurobiology of Disease, 59C: 257-266. PMCID: PMC3790640.

d. Goffin D, Brodkin ES, Blendy JA, Siegel SJ and Zhou Z* (2014). Cellular origins of auditory event-related potential deficits in Rett syndrome. Nature Neuroscience, 17(6): 804-806. PMCID: PMC4038660.

2. The field of Rett syndrome research has been greatly benefited and advanced through studies of knockout, conditional knockout and conditional rescue mouse models of MeCP2. However, nearly one-third of Rett syndrome mutations are missense in the methyl-CpG binding domain (MBD) of MeCP2. To specifically address the contribution of these missense mutations to the pathogenesis of Rett Syndrome and provide the research community with clinically relevant mouse models, I led a research team and developed the first allelic series of knockin models that faithfully recapitulate patient mutations, MeCP2 T158A, T158M and R106W. We found that those missense mutations impair the binding of MeCP2 to methylated DNA in vivo, concomitantly reduce MeCP2 protein stability, and lead to behavioral phenotypes resembling Rett syndrome. Notably, we uncovered that stabilizing MeCP2 T158M protein or elevating MeCP2 T158M expression rescues Rett-like phenotypes in vivo, providing a novel therapeutic strategy to treat Rett syndrome. Though studies of these mouse models, we also discovered that event-related neuronal responses are stalled in maturation by the loss-of-function of Mecp2, and auditory ERPs represent robust, sensitive, and quantitative biomarkers informing phenotypic progression, at least in mouse models. Furthermore, we demonstrated both cell and non-cell autonomous effects of MeCP2 on gene expression and the existence of post-transcriptional compensation to MeCP2 loss. These findings allowed us to postulate that MeCP2 functions as an architectural protein organizing chromatin in 3-dimentional fashion to facilitate precise gene transcription.

a. Goffin D, Allen M, Amorim M, Zhang L, Wang I-TJ, Reyes A-RS, Mercado-Berton A, Ong C, Cohen S, Hu L, Blendy JA, Carlson G,Siegel S, Greenberg ME and Zhou Z* (2012). Rett Syndrome mutation MeCP2 T158A mutation disrupts DNA binding, protein stability and ERP responses. Nature Neuroscience, 15: 274-283. PMCID: PMC3267879.

b. Lamonica JM, Kwon DY, Goffin D, Fenik P, Johnson BS, Cui Y, Guo H, Veasey S and Zhou Z* (2017). Elevating expression of MeCP2 T158M rescues DNA binding and Rett syndrome-like phenotypes. Journal of Clinical Investigation, 127 (5): 1889-1904. PMCID: PMC5409785.

c. Johnson BS, Zhao Y, Fasolino M, Lamonica JM, Kim YJ, Georgakilas G, Wood KH, Bu D, Cui Y, Goffin D, Vahedi G, Kim TH and Zhou Z* (2017). Biotin tagging of MeCP2 reveals contextual insights into the Rett syndrome transcriptome. Nature Medicine, 23(10): 1203-1214. PMCID: PMC5630512.

d. Connolly DR, Zhou Z* (2019). Genomic insights into MeCP2 function: A role for the maintenance of chromatin architecture. Current Opinion in Neurobiology, 59:174-179. PMCID: PMC6889049.

3. While I was in the process of studying MeCP2 phosphorylation and trying to identify its up-stream kinase, several human genetic studies linked mutations in the X-linked gene encoding cyclin-dependent kinase-like 5 (CDKL5) to atypical Rett syndrome, a variant with early-onset epileptic features. While several in vitro biochemistry and cell culture studies began to support an interaction between CDKL5 and MeCP2, the experimental evidence remained contentious. I decided to take a genetic approach to investigate CDKL5 function in vivo, and therefore, led a research team to the development and characterization of the first knockout mouse model of CDKL5 deficiency disorder (CDD). We found that CDKL5 dysfunction disrupts many key signal transduction pathways and long-range neural circuit communication, leading to autistic-like phenotypes in mice. We have also carried out conditional knockout studies to dissect the cellular origin and uncovered a crucial role for CDKL5 in glutamatergic neurons for learning and memory, in GABAergic neurons for social interaction and repetitive behaviors. Through temporal manipulation of CDKL5 expression in vivo, we uncovered that CDKL5 is essential for post-developmental brain function, and importantly, CDD-related behavioral and synaptic phenotypes are reversible after the early stage of brain development. We have now developed and contributed multiple mouse models of CDD to the community, demonstrated the potential for symptomatic reversal, at least in mouse models, and paved the way for mechanistic and therapeutic studies of CDD.

a. Wang IT, Allen M, Goffin D, Zhu X, Fairless AH, Brodkin ES, Siegel SJ, Marsh ED, Blendy JA, and Zhou Z* (2012). Loss of CDKL5 disrupts kinome profile and ERP response leading to autistic-like phenotypes in mice. Proc Natl Acad Sci USA.109: 21516-21521. PMCID: PMC3535652.

b. Tang S, Wang I-T, Yue C, Takano H, Terzic B, Pance K, Lee JY, Cui Y, Coulter DA* and Zhou Z* (2017). Loss of CDKL5 in glutamatergic neurons disrupts hippocampal microcircuitry and leads to memory impairment in mice. Journal of Neuroscience, 37(31): 7420-7437. PMCID: PMC5546111.

c. Terzic B, Cui Y, Edmondson AC, Tang S, Sarmiento N, Zaitseva D, Marsh ED, Coulter DA and Zhou Z* (2021): X-linked cellular mosaicism underlies age-dependent occurrence of seizure-like events in mouse models of CDKL5 deficiency disorder. Neurobiology of Disease, 148: 105176. doi: 10.1016/j.nbd.2020. 105176. PMCID: PMC7856307.

d. Terzic B, Davatolhagh MF, Ho Y, Tang S, Liu Y-T, Xia Z, Cui Y, Fuccillo MV and Zhou Z* (2021): Temporal manipulation of Cdkl5 reveals essential post-developmental functions and reversible CDKL5 deficiency disorder-related deficits. Journal of Clinical Investigation, 131(20): e143655, 2021. PMCID: PMC8516470.

4. The genetic underpinnings of mental health disorders are highly complex, involving multifaceted interactions between risk genes and the environment. It is known that environmental factors such as adverse early life events confer significantly greater susceptibility to psychiatric conditions later in life. But, the epigenetic mechanisms by which environmental factors interact with genetic programs in the nervous system remain poorly understood. Sponsored by the Biobehavioral Research Award for Innovative Scientist (BRAINS) from NIMH, I led a research team conceived a novel Cre-dependent biotinylation strategy and developed a series of genetically modified mice that allow for genome wide profiling of DNA methylation, histone modifications and RNA expression from cell types of interest while overcoming extensive cellular heterogeneity in the brain. We found that long genes genetically implicated in autism harbor broad enhancer-like chromatin domains (BELD) and causally link chromatin genes to autism etiology. My research team have also developed a computational pipeline to identify integrated epigenetic code in cell types of interest and in response to environmental stimuli. To investigate the causal role of stress-induced epigenetic changes to behavioral maladaptation, my research team have also adopted a modified CRISPR/Cas9 strategy to alter DNA methylation and histone acetylation at loci of interest. Notably, we recently found that chronic stress impacts higher-order chromatin architecture and identified YY1 as a regulator of stress-induced maladaptive behavior, thus providing a molecular target that mediates gene-environment interactions in the context of stress-related neuropsychiatric conditions.

a. Wood KH, Johnson BS, Welsh SA, Lee JY, Cui Y, Krizman E, Brodkin ES, Blendy JA, Robinson MB, Bartolomei MS and Zhou Z* (2016): Tagging of Methyl-CpG-binding Domain Proteins Reveals Different Spatiotemporal Expression and Supports Distinct Functions. Epigenomics, 4: 455-473. PMCID: PMC4880565.

b. Kwon DY, Zhao YT, Lamonica JM and Zhou Z* (2017). Locus-specific histone deacetylation using a synthetic CRISPR-Cas9-based HDAC. Nature Communications, 8:15315. Doi:10.1038/ncomms15315. PMCID: PMC5437308.

c. Zhao YT, Kwon DY, Johnson BS, Fasolino M, Lamonica JM, Kim YJ, Zhao BS, He C, Vahedi G, Kim TH and Zhou Z* (2018). Long genes linked to autism harbor broad enhancer-like chromatin domains. Genome Research, 28:933-942. PMCID: PMC6028126.

d. Kwon DY, Xu B, Hu P, Zhao Y-T, Beagan JA, Nofziger JH, Cui Y, Phillips-Cremins JE, Blendy JA, Wu H, Zhou Z* (2022): Neuronal YY1 in the prefrontal cortex regulates transcriptional and behavioral responses to chronic stress. Nature Communications, 13: 55. doi.org/10.1038/s41467-021-27571-3. PMCID: PMC8748737.