Characterising pre-MBT nucleus and transcriptional dynamics in the developing zebrafish embryo

Qureshi, Haseeb Khalid (2023). Characterising pre-MBT nucleus and transcriptional dynamics in the developing zebrafish embryo. University of Birmingham. Ph.D.

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Abstract

The early embryo begins life in a transcriptionally silent state, and undergoes a process of genomic activation as it is reprogrammed from an egg cell into a mass of stem cells. In the zebrafish, this process of genome awakening, the zygotic genome activation, occurs over several embryonic stages and consists of rapid cell cycles which exclusively switch between S-phase and mitosis. This raises the question of how genome awakeningisabletooccurconcurrentlywithagenomeoccupiedbyDNAreplicationevents. We investigated how transcription is coordinated with the cell cycle. Through developing image processing and analysis tools, I was able to parse complex 4D microscopy datasets to characterise the progressive loss of mitotic synchronyandincreaseincellcyclelengthsinthephasespriortomidblastulatransition(MBT).3Dvisualisation and analysis of high resolution imaging of microRNA-430 morpholino visualised transcription revealed large localised transcription compartments within nuclei, termed here as ’foci’, which contain transcripts from multiple genes. The presence and duration of transcription occurs a coordinated manner, showing heritable relationships between parent and daughter cells, and the cell cycle duration showed a similar relationship. This provoked a question regarding how cell cycle influences transcription as a heritable feature. These analyses also revealed the limitations of conventional image processing methods, so I designed a 4D microscopy image analysis tool which utilised a novel iteration of the watershed algorithm to segment nuclei in highly noisy deep tissue environments. This tool was also able to trace nuclei over generations to provide lineages, with data about cell cycles and nuclear geometries. Based off this, I was able to make early characterisations of potential cell cycle memory effects between parent and daughter cells in pre-MBT cell cycles. I also analysed these variations in cell cycle lengths in the context of nucleocytoplasmic ratio, and position in the embryo, shedding light on how cell cycle variation and asynchrony can influence a mitotic wave, which spreads across the embryo.

Type of Work:

Thesis (Doctorates > Ph.D.)

Award Type:

Doctorates > Ph.D.

Supervisor(s):