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During development, the hypothalamus emerges from the ventral diencephalon and is regionalized into several distinct functional domains. Each domain is characterized by a different combination of transcription factors, including Nkx2.1, Nkx2.2, Pax6, and Rx, which are expressed in the presumptive hypothalamus and its surrounding regions, and play critical roles in defining each area. Here, we recapitulated the molecular networks formed by the gradient of Sonic Hedgehog (Shh) and the aforementioned transcription factors. Using combinatorial experimental systems of directed neural differentiation of mouse embryonic stem (ES) cells, as well as a reporter mouse line and gene overexpression in chick embryos, we deciphered the regulation of transcription factors by different Shh signal intensities. We then used CRISPR/Cas9 mutagenesis to demonstrate the mutual repression between Nkx2.1 and Nkx2.2 in a cell-autonomous manner; however, they induce each other in a non-cell-autonomous manner. Moreover, Rx resides upstream of all these transcription factors and determines the location of the hypothalamic region. Our findings suggest that Shh signaling and its downstream transcription network are required for hypothalamic regionalization and establishment., (© The Author(s) 2023. Published by Oxford University Press. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Methylated DNA can be specifically recognized by a set of proteins called methyl-CpG-binding proteins (MBPs), which belong to three different structural families in mammals: the MBD family, the Kaiso and Kaiso-like proteins and the SRA domain proteins. A current view is that, once bound to methylated DNA, MBPs translate the DNA methylation signal into appropriate functional states, through interactions with diverse partners. However, if some of the biological functions of MBPs have been widely described--notably transcriptional repression--others are poorly understood, and more generally the extent of MBP activities remains unclear. Here we propose to discuss the role of MBPs in two crucial nuclear events: chromatin organization and epigenome maintenance. Finally, important challenges for future research as well as for biomedical applications in pathologies such as cancers--in which DNA methylation patterns are widely altered--will be mentioned.

DNA methylation is an essential epigenetic mark. Three classes of mammalian proteins recognize methylated DNA: MBD proteins, SRA proteins and the zinc-finger proteins Kaiso, ZBTB4 and ZBTB38. The last three proteins can bind either methylated DNA or unmethylated consensus sequences; how this is achieved is largely unclear. Here, we report that the human zinc-finger proteins Kaiso, ZBTB4 and ZBTB38 can bind methylated DNA in a sequence-specific manner, and that they may use a mode of binding common to other zinc-finger proteins. This suggests that many other sequence-specific methyl binding proteins may exist.

A replication initiator protein (RepE54) complexed with iteron DNA at its binding site was crystallized by the hanging drop vapor diffusion method. The crystals belong to monoclinic space group C2 with unit cell dimensions of a = 108.4 A, b = 81.9 A, c = 73.9 A, and beta = 121.5 degrees, where one molecule of the protein-DNA complex exists per asymmetric unit. They diffract X-rays up to 2.6 A resolution with synchrotron radiation.