Biology subjects

Rusimbi, C.

Publications and source records attributed to Rusimbi, C..

3 recordsLinked to original sources

bioRxiv2025

Mapping early patterning events in human neural development usingan in-vitro microfluidic stem cell model

Stem cell models can provide insights into human brain development at embryonic stages which are normally inaccessible. We previously developed the Microfluidic Stem Cell Regionalisation (MiSTR) model, which recapitulates the rostro-caudal patterning of human neural tube through a WNT activation (WNTa) gradient. Through temporal single cell transcriptomics of rostro-caudal and dorso-ventral gradient-patterned MiSTR, we found that rostro-caudal subtypes were regionally specified and fate-determined already during the late epiblast stage, several days before onset of neuralisation at day 3-4. Rostral cells were characterised by expression of HESX1 and SHISA2 during pre-neuralisation and PAX6 during early neuralisation, whereas caudal cells expressed FST and HOXA1 during pre-neuralisation and SOX1 as the dominant neuralising factor. In contrast to the early rostro-caudal specification, response to ventralisation in telencephalic progenitors was developmentally delayed and occurred around day 9. We further uncovered temporal events in human midbrain-hindbrain boundary formation and ventral forebrain patterning, contributing new knowledge on early human neural region-specification.

neuroscience↗

bioRxiv2024

PAF1C allosterically activates CDK12/13 kinase during RNAPII transcript elongation

The mechanisms ensuring temporally correct, site-specific phosphorylation of the RNA polymerase II C-terminal domain (CTD) by cyclin-dependent kinases (CDKs) during the transcription cycle remain poorly understood. Here, we present results from in vitro reconstitution of CTD phosphorylation combined with in vivo evidence to show that human CDK12 and CDK9 both co-phosphorylate CTD Serine 5 and Serine 2. However, only phosphorylation by CDK12 is stimulated by association with the elongation-specific factor PAF1C, in which the CDC73 subunit contains a short, conserved motif capable of association with and activation of CDK12/Cyclin K. This motif is necessary for cell proliferation and crucial for CTD phosphorylation and transcript elongation. Together, these data provide new insight into basic mechanisms ensuring CDK specificity in the RNAPII transcription cycle. One-Sentence SummaryPAF1C facilitates RNAPII phosphorylation in gene bodies through direct contacts with the active site of CDK12/13.

molecular biology↗

bioRxiv2024

Decoding morphogen patterning of human neural organoids with a multiplexed single-cell transcriptomic screen

Morphogens, secreted signalling molecules that direct cell fate and tissue development, are used to direct neuroepithelial progenitors towards discrete regional identities across the central nervous system. Neural tissues derived from pluripotent stem cells in vitro (neural organoids) provide new models for studying neural regionalization, however, we lack a comprehensive survey of how the developing human neuroepithelium responds to morphogen cues. Here, we produce a detailed map of morphogen-induced effects on the axial and regional specification of human neural organoids using a multiplexed single-cell transcriptomics screen. We find that the timing, concentration, and combination of morphogens strongly influence organoid cell type and regional composition, and that cell line and neural induction method strongly impact the response to a given morphogen condition. We apply concentration gradients in microfluidic chips or a range of static concentrations in multi-well plates to explore how human neuroepithelium interprets morphogen concentrations and observe similar dose-dependent induction of patterned domains in both scenarios. Altogether, we provide a detailed resource that supports the development of new regionalized neural organoid protocols and enhances our understanding of human central nervous system patterning.

developmental biology↗