Trp53 ablation fails to prevent microcephaly in mouse pallium with impaired minor intron splicing

Abstract

Minor spliceosome inhibition due to mutations in RNU4ATAC are linked to primary microcephaly. Ablation of Rnu11, a minor spliceosome snRNA, inhibits the minor spliceosome in the developing mouse pallium, causing microcephaly. There, cell cycle defects and p53-mediated apoptosis in response to DNA damage resulted in loss of radial glial cells (RGCs), underpinning microcephaly. Here, we ablated Trp53 to block cell death in the Rnu11 cKO mice. We report that Trp53 ablation failed to prevent microcephaly in these double knockout (dKO) mice. We show that the transcriptome of the dKO pallium was closer to the control compared to the Rnu11 cKO. We find aberrant minor intron splicing in MIGs involved in cell cycle regulation, resulting in more severely impaired mitotic progression and cell cycle lengthening of RGCs in the dKO that was detected earlier than the Rnu11 cKO. Furthermore, we discover a potential role of p53 in causing DNA damage in the developing pallium, as detection of γH2aX+ was delayed in the dKO. Thus, we postulate that microcephaly in minor spliceosome-related diseases is primarily caused by cell cycle defects.

Integrity of the minor spliceosome in the developing mouse hypothalamus determines neuronal subtype composition regulating energy balance

“…Here, we show that integrity of the minor spliceosome is essential for hypothalamic development and function. Specifically, we show that ablation of Rnu11 inhibits the minor spliceosome, which resulted in elevated minor intron retention and aberrant AS of MIGs involved in cell cycle regulation, consistent with previous reports 40,51,44,43 . As expected, we found progenitor cell cycle defect in the VD ( Fig.…”

“…To assess minor intron splicing defects, we next performed bulk RNA-seq on microdissected E12.5 tdTomato+ VD for control (n=3) and mutant (n=3) embryos (Fig. 1D) 40,43,44 . The expression of Nkx2.1, Six3, and Rax combined with the absence of Emx1 and Foxg1, confirmed the accuracy of our dissection [45][46][47] (Supplementary Data Table 1).…”

“…To assess minor intron splicing defects, we next performed bulk RNA-seq on microdissected E12.5 tdTomato+ VD for control (n=3) and mutant (n=3) embryos ( Fig. 1D ) 40,43,44 . The expression of Nkx2 .…”

Integrated multi-omics reveals minor spliceosome inhibition causes molecular stalling and developmental delay of the mouse forelimb

“…11). Future experiments leveraging ablation of p53 (to block LPC apoptosis) in the U11-null forelimb would help resolve primary phenotypes occurring from mis-splicing of MIGs and secondary phenotypes resulting from cell death 69 . Nonetheless, our systems analysis reveals that the minor spliceosome is required not only for the proper development of endogenous limb progenitors, but also to generate the molecular framework from which the entire limb will be constructed (Fig.…”

P53 independent pathogenic mechanisms contribute to BubR1 microcephaly

“…P53 activation can arrest the cell cycle or initiate apoptosis to eliminate genomically unfit cells as part of its role in mitotic surveillance (Pilaz et al, 2016;Soto et al, 2017;Hafner et al, 2019;Navarro et al, 2020). Importantly, P53 activation has been observed in a variety of microcephaly models (Frank et al, 2000;Gao, et al, 2000;McConnell et al, 2004;Insolera et al, 2014;Marjanovic et al, 2015;Ghouzzi et al, 2016;Mao et al, 2016;Bianchi et al, 2017;Sgourdou et al, 2017;Little and Dwyer, 2019;Shi et al, 2019;Phan et al, 2021;White et al, 2021;Martins et al, 2022;Peek et al, 2022;Sterling et al, 2023).…”