The role of chromatin modifications in the efficiency of reprogramming
General Genetics and Functional Genome Biology
Final Report Abstract
We investigated the role of H3K4 methyltransferases and H3K27 demethylases in embryonic development, ESC pluripotency and reprogramming to iPSCs. We found that Mll1 regulates neural specific genes in NSCs and its conditional inactivation facilitates reprogramming to iPSCs, In contrast Mll2, the paralog of Mll1, has an opposite effect in induced reprogramming efficiency. These observations together with the different embryonic and adult phenotypes of Mll1 and Mll2 knockout, strongly argue that the two methyltransferases regulate distinct target genes. Similar results were obtained with the Setd1a and Setd1b paralogs. Despite highly homologous protein sequences and identical protein complexes, Setd1a and Setd1b play very different essential roles in mouse development. Setd1a is required immediately before gastrulation shortly after implantation. At this stage and in ESCs, it is the major H3K4 methyltransferase. Setd1a is also essential in ESCs for proliferation and maintenance of the self-renewal circuitry. However, loss of this circuitry in Setd1a knockout ESCs did not provoke default differentiation suggesting that Setd1a may be required for ESC transcriptional activity in general, which is a proposition supported by the fact that it is the major ESC H3K4 methyltransferase. These observations are reinforced by our reprogramming experiments from NSCs. Setd1a is required for NSC proliferation and its removal at any stage during a reprogramming time course impeded the acquisition of iPSC lines. In contrast, none of these functions apply to Setd1b, which is required after gastrulation during organogenesis and its deletion has no effect in ESC pluripotency maintenance or reprogramming to iPSCs. We also found that the H3K27 demethylase Utx plays a role in reprogramming efficiency. Conditional deletion of Utx impairs reprogramming whereas inducible Utx expression increases the reprogramming efficiency. There is little known about the direct targets of HMTs and histone demethylases in different cell types. The experimental systems that we have established together with genome wide RNA-seq and ChIP-seq analysis will contribute in answering these fascinating questions in the near future.
Publications
- (2009): A genome-scale RNAi screen for Oct4 modulators defines a role of the Paf1 complex for embryonic stem cell identity. Cell Stem Cell 4(5): 403-15
Ding L, Paszkowski-Rogacz M, Nitzsche A, Slabicki MM, Heninger AK, de Vries I, Kittler R, Junqueira M, Shevchenko A, Schulz H, Hubner N, Doss MX, Sachinidis A, Hescheler J, Iacone R, Anastassiadis K, Stewart AF, Pisabarro MT, Caldarelli A, Poser I, Theis M, Buchholz F
(See online at https://doi.org/10.1016/j.stem.2009.03.009) - (2010): Mll2 is required in oocytes for bulk histone 3 lysine 4 trimethylation and global transcriptional silencing. PLoS Biology 8(8): e1000453
Andreu-Vieyra CV, Chen R, Agno J, Glaser S, Anastassiadis K, Stewart AF and Matzuk MM
(See online at https://doi.org/10.1371/journal.pbio.1000453) - (2013): A localized Wnt signal orients asymmetric stem cell division in vitro. Science 339(6126): 1445-8
Habib SJ, Chen BC, Tsai FC, Anastassiadis K, Meyer T, Betzig E, Nusse R
(See online at https://doi.org/10.1126/science.1231077) - (2013): Biallelic expression of nanog protein in mouse embryonic stem cells. Cell Stem Cell 13(1): 12-3
Filipczyk A, Gkatzis K, Fu J, Hoppe PS, Lickert H, Anastassiadis K, Schroeder T
(See online at https://doi.org/10.1016/j.stem.2013.04.025) - (2013): The histone demethylase UTX regulates stem cell migration and hematopoiesis. Blood. 121(13): 2462-73
Thieme S, Gyárfás T, Richter C, Ozhan G, Fu J, Alexopulou D, Muders MH, Michalk I, Jakob C, Dahl A, Klink B, Bandola J, Bachmann M, Schröck E, Buchholz F, Stewart AF, Weidinger G, Anastassiadis K, Brenner S
(See online at https://doi.org/10.1182/blood-2012-08-452003) - (2014): Mll2 is required for H3K4 trimethylation on bivalent promoters in embryonic stem cells, whereas Mll1 is redundant. Development 141(3): 526-37
Denissov S, Hofemeister H, Marks H, Kranz A, Ciotta G, Singh S, Anastassiadis K, Stunnenberg HG, Stewart AF
(See online at https://doi.org/10.1242/dev.102681) - (2014): The H3K4 methyltransferase Setd1a is first required at the epiblast stage, whereas Setd1b becomes essential after gastrulation. Development 141(5): 1022-35
Bledau AS, Schmidt K, Neumann K, Hill U, Ciotta G, Gupta A, Torres DC, Fu J, Kranz A, Stewart AF, Anastassiadis K
(See online at https://doi.org/10.1242/dev.098152)
