Biological Session II

BASIC NEUROSCIENCE

Regulatory genomics of astrocyte evolution

Ciuba K.1 Piotrowska A. 1, Chaudhury D. 1, Dehingia B.1, Dunski E.1, Wójtowicz T.2, Włodarczyk J.2, Aleksandra Pękowska1,2

1 Dioscuri Centre for Chromatin Biology and Epigenomics, Nencki Institute of Experimental Biology, Polish Academy of Sciences
2 Laboratory of Cell Biophysics, Nencki Institute of Experimental Biology, Polish Academy of Sciences, Poland

Abstract: Astrocytes play an essential role in the maintenance of brain homeostasis. Virtually all neurological disorders are related to astrocyte dysfunctions. Remarkably, astrocytes have changed profoundly during the evolution of the mammalian brain. Human brains feature at least four morphotypes of astrocytes, while only one is present in the mouse brain. Furthermore, the evolution of primate astrocytes led to an enhanced complexity of protoplasmic astrocyte arborization. Recent transcriptomic comparisons of human, chimpanzee, and macaque cortical cells suggest that astrocytes might have evolved faster than neuronal cells. Molecular bases, as well as the functional significance of these changes, remain vastly unexplored. We used induced pluripotent stem cells to obtain human, chimpanzee, and macaque astrocytes in vitro. In my presentation, I will summarize our data highlighting the functional classes of genes significantly deregulated in human astrocytes compared to chimpanzees and macaque cells. I will discuss the relevance of these loci to human diseases. I will outline the gene and transcriptional regulatory network changes accompanying astrocyte evolution and give numerous examples of how human-specific enhancer elements might account for the differences in gene expression levels in cells of our species. I will discuss the epigenetic signature of promoter elements in astrocyte evolution. Our data will help better understand astrocytes' implications in brain evolution and shed new light on the link between human-specific gene expression and neurological disorders.

Evolutionary features of human astrocytes

Katarzyna Ciuba, Aleksandra Piotrowska, Eryk Duński, Debadeep Chaudhury,  Bondita Dehingia, Aleksandra Pękowska

Dioscuri Center for Chromatin Biology and Epigenomics, Nencki Institute of Experimental Biology, Polish Academy of Sciences, Warsaw, Poland

Abstract: Cognitive ability of human brain is exceptional comparing to other species, including non-human primates. However, its genetic basis remains one of the biggest unsolved questions of modern biology. To date, extensive research focused on neuronal cells. Yet, recent advancements in the field indicated the importance of astrocyte participation in regulation of neuronal plasticity and consequently cognitive processes. Importantly, astrocyte properties have changed remarkably throughout evolution. Human astrocytes are bigger and more complex than rodent ones, pointing to an evolutionary interplay between astrocyte properties and cognition. Nonetheless, genetic and functional determinants of astrocyte biology remain largely unknown.  

To identify evolutionary changes, driving human-specific astrocyte expansion, we established an in vitro model of iAstrocytes (iPS-derived astrocytes) from distinct primate species (H. sapiens, P. Troglodytes, M. Mulatta). By combining molecular biology, high-throughput genomics, including RNA-Seq, ATAC-Seq, ChIP-Seq, and bioinformatic methods, we identified genes and gene regulatory elements featuring differential activity between human and other primate species. We apply CRISPR/Cas9-based methods to examine function of particular, human specific, up- or down-regulated loci. This new data reveals human-specific genes and epigenetic regulatory networks. Consequently, our results will be instrumental for understanding the nature of the contribution of evolutionary changes in astrocyte epigenome and human specific brain functions.

Funding: Dioscuri Grant. Dioscuri is a program initiated by the Max Planck Society, managed jointly with the National Science Centre (Poland) and mutually funded by the Polish Ministry of Science and Higher Education and the German Federal Ministry of Education and Research. 

CRISPR/Cas9-mediated ablation of full-length TCF7L2 alters  thalamic phenotype in developing mice

Michael Gabriel, Marcin Lipiec, Joanna Bem, Kacper Posyniak,  Marta Wiśniewska

Laboratory of Molecular Neurobiology, Centre of New Technologies, University of Warsaw, Warsaw, Poland.

Abstract: The Wnt/β-catenin pathway is an evolutionarily conserved signalling pathway that regulates many biological processes during embryonic development. Transcription factor-7 like 2 (TCF7L2) is an effector of the Wnt/β-catenin signalling pathway that plays different roles in the development of the thalamus. We show that two groups of TCF7L2 isoforms are ubiquitously expressed in the thalamus at different developmental stages: the full-length (fl-TCF7L2) and truncated (dn-TCF7L2) isoforms. Although these isoforms differ respectively by the presence and absence of the β-catenin binding domain, studies have suggested that both of them act independently of β-catenin. The distinct roles of these isoforms in thalamic development remain enigmatic. We used CRISPR/Cas9 technology to specifically knockout fl-TCF7L2 by targeting exon 2. The expression of the dn-TCF7L2 isoform was preserved. We demonstrate that fl-TCF7L2 is vital for cell sorting and boundary formation during early thalamic development. We also show that fl-TCF7L2 regulates the expression of sub-regional thalamic markers highlighting its role in assigning thalamus-specific features to neurons. However, the fl-TCF7L2 knockout did not affect the guiding of thalamic axons into the ventral telencephalon. Overall, this study provides a robust understanding of the in vivo regulatory activities of fl-TCF7L2 on the development of the thalamus.

Funding:  This project is supported by the National Science Centre in Poland. (Grant No. UMO-2017/25/B/NZ3/01665)

Mathematical and experimental understanding of the mechanisms associated with the fluid excitability and “flipping” of maturating granule cells

Danielewicz J.1,2, Girier G.2, Chizhov A.3,4, Desroches M.3, Encinas JM1,2,3, Rodrigues S.2,6

1Achucarro Basque Center for Neuroscience, Leioa, Bizkaia,Spain  
2BCAM Basque Center for Applied Mathematics, Bilbao, Bizkaia, Spain  
3Inria Sophia Antipolis - Méditerranée Research Centre, Paris, France  
4 Ioffe Physical Technical Institute, Saint Petersburg, St.-Petersburg, Russia  
5 University of the Basque Country (UPV/EHU), Leioa, Bizkaia, Spain  
6Ikerbasque, The Basque Science Foundation, Bilbao, Bizkaia, Spain 

Abstract: In response to prolonged depolarizing current steps, different classes of neurons display specific firing characteristics (i.e. excitability class), such as a regular train of action potentials with more or less adaptation, delayed responses, or bursting. In general, one or more specific ionic transmembrane currents underlie the different firing patterns. The intrinsic firing properties and ionic conductances in granule cells (GCs) are thought to reflect their developmental stage and maturation level. Among GCs, input resistance, threshold current, fluid excitability and the transition to depolarization block (DB) have been used as signatures of the degree of maturation and circuitry integration. Here we sought to investigate the influence of sodium and potassium channels conductances on DB in GCs with dynamic clamp – a computer controlled real-time closed-loop electrophysiological technique, which allows to couple mathematical models simulated in a computer with biological cells. We have observed that 44% of recorded cells exhibited what we have called a “flipping” behavior. Meaning, that these cells were able to overcome the DB and generate trains of action potentials at later current injections steps. We have develop a unified mathematical model of maturating GCs to explain fluid excitability, “flipping” and to capture the essential features of entry into DB.

Funding:  This project received support from Ikerbasque (The Basque Foundation for Science), the Basque Government through the BERC 2018-2021 and BERC 2022-2025 program, PIBA_2021_1_0018 and MICINN PID2019-104766RB-C21, the Spanish State Research Agency through BCAM Severo Ochoa excellence accreditation SEV-2017-0718, “MathNEURO” project RTI2018-093860-B-C21 funded by (AEI/FEDER, UE). Moreover, we acknowledge the support of Inria via the Associated Team “NeuroTransSF”.

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