Shajahan, S. | bioRxiv Science

bioRxiv2025

Z-DNA formation induces the totipotent-like state and primes Zscan4-dependent chromatin compartmentalization

A remarkable transition during murine development is the progression from the 1-cell embryo to the 2-cell stage, accompanied by the activation of a specific set of embryonic genes, epigenome reprogramming, and nuclear architecture reorganization. Some of these characteristics are recapitulated in vitro with the spontaneous emergence of 2-cell-like cells from mouse embryonic stem cells, which exhibit a transcriptomic signature resembling the 2-cell stage, including the expression of genes such as Dux, Zscan4, and the repetitive element MERVL, and a more relaxed chromatin state. Here, we show that inter- and intra-chromosomal interactions, driven by Zscan4 chromatin factors, form during this transition and segregate into a distinct genomic compartment (Z compartment). Mechanistically, the formation of Z-DNA, an alternative DNA conformation regulated by polyamine levels, promotes the emergence of totipotent-like cells and the establishment of the Z compartment. This compartment is characterized by a decrease in active histone marks and a reduced expression of genes associated with differentiation and late developmental processes. Overall, these findings suggest that Z-DNA formation may play a dual role, first in activating ZGA genes and later in guiding genome compartmentalization to safeguard the totipotent-like state by restricting the expression of non-ZGA genes within a permissive chromatin environment.

cell biology↗

bioRxiv2022

Increased Relative Biological Effectiveness of Orthovoltage X-rays Compared to γ-rays in Preclinical Irradiation

PurposeRadionuclide irradiators (137Cs and 60Co) are commonly used in preclinical studies ranging from cancer therapy to stem cell biology. There are institutional initiatives to replace radionuclide sources with lower-energy X-ray sources amidst concerns of radiological terrorism. As researchers transition, there are questions whether the biological effects of {gamma}-rays may be recapitulated with orthovoltage X-rays, since different energy may cause different biological effects. We, therefore, sought to compare the effects of orthovoltage X-rays and 137Cs {gamma}-rays using mouse models of acute radiation syndrome. Experimental Design137Cs {gamma}-rays were compared with Orthovoltage X-rays, generated at 300 kVp, 10 mA with 1 mm Cu or Thoraeus filtration. We assessed 30-day overall survival following whole-body irradiation and calculated LD50 by logistic regression. Comparing equivalent doses delivered with different average energies ([E]), we assessed bone marrow, spleen, and intestinal histology and flow cytometry. ResultsThe LD50 doses are 6.7 Gy, 7.4 Gy and 8.1 Gy with 1 mm Cu filtered ([E]=120 keV), and Thoraeus filtered X-rays ([E]=160 keV), and 137Cs (E=662 keV), respectively. At constant dose, hematopoietic injury was most severe with 1 mm Cu filtered X-rays with the greatest reduction in bone marrow cellularity, stem and progenitor populations, and intestinal crypts and OLFM4+ intestinal stem cells. Thoraeus filtered X-rays provoked an intermediate phenotype, with 137Cs showing the least damage. ConclusionsOur study reveals a dichotomy between physical dose and biological effect relevant as researchers transition to orthovoltage X-rays. With decreasing energy, there is increasing hematopoietic and intestinal injury, necessitating dose-reduction to achieve comparable biological effects. Statement of Translational RelevanceRadiation is used in translational studies in fields ranging from hematopoiesis and stem cell biology to cancer radiotherapy, with 137Cs and 60Co radionuclide sources serving as the most common irradiators. Due to the threat of radiological terrorism using stolen radionuclides, there are institutional initiatives to replace these sources with orthovoltage X-ray irradiators. Yet, as shown in this study, the biological effects of radiation are highly dependent on radiation energy. Lower energy orthovoltage X-rays are absorbed differently than higher energy radionuclide {gamma}-rays, provoking more severe hematopoietic, immunologic, and gastrointestinal radiation injury. Thus, an identical physical dose delivered with beams of differing energy does not produce the same biologic effect. As researchers transition between these sources, it is critical that we appreciate that radiation doses are not interchangeable between them. Understanding the significance of physical dose delivered using different methods will allow us to contextualize past results with future studies.

pathology↗