Biology subjects

Bahiru, M. S.

Publications and source records attributed to Bahiru, M. S..

3 recordsLinked to original sources

bioRxiv2025

Lamin B loss in nuclear blebs is rupture dependent while increased DNA damage is rupture independent

The nucleus houses genetic information and functions separate from the rest of the cell. Loss of nuclear shape results in nuclear ruptures. Nuclear blebs are deformations identified by decreased DNA density, while lamin B levels vary drastically. To determine if decreased lamin B levels are due to nuclear rupture, we used immunofluorescence to measure levels of lamin B and emerin, a nuclear envelope protein that enriches to sites of nuclear rupture. We observed that cell types that exhibit decreased levels of lamin B also show an enrichment of emerin in nuclear blebs. Oppositely, in other cell types, nuclear blebs display maintained levels of lamin B1 and showed no emerin enrichment. To determine how nuclear rupture affects DNA damage, we time lapse imaged nuclear rupture dynamics then fixed the same cells to conduct immunofluorescence of {gamma}H2AX and emerin. We find that DNA damage levels are higher in blebbed nuclei independent of nuclear rupture. Thus, we confirm that lamin B1 loss in nuclear blebs is due to nuclear rupture and blebbed nuclei have increased DNA damage that is independent of rupture. Summary statement (180-200 characters): We measured lamin B and DNA damage in blebbed nuclei to determine the effect of nuclear rupture. We find that nuclear rupture causes loss of lamin B in nuclear blebs but that increased DNA damage in blebbed nuclei is independent of rupture.

cell biology↗

bioRxiv2024

DNA density is a better indicator of a nuclear bleb than lamin B loss

Nuclear blebs are herniations of the nucleus that occur in diseased nuclei that cause nuclear rupture leading to cellular dysfunction. Chromatin and lamins are two of the major structural components of the nucleus that maintain its shape and function, but their relative roles in nuclear blebbing remain elusive. Lamin B is reported to be lost in blebs by qualitative data while quantitative studies reveal a spectrum of lamin B levels in nuclear blebs dependent on perturbation and cell type. Chromatin has been reported to be decreased or de-compacted in nuclear blebs, but again the data are not conclusive. To determine the composition of nuclear blebs, we compared the immunofluorescence intensity of lamin B and DNA in the main nucleus body and nuclear bleb across cell types and perturbations. Lamin B nuclear bleb levels varied drastically across MEF wild type and chromatin or lamins perturbations, HCT116 lamin B1-GFP imaging, and human disease model cells of progeria and prostate cancer. However, DNA concentration was consistently decreased to about half that of the main nucleus body across all measured conditions. Using Partial Wave Spectroscopic (PWS) microscopy to measure chromatin density in the nuclear bleb vs body we find similar results that DNA is consistently less dense in nuclear blebs. Thus, our data spanning many different cell types and perturbations supports that decreased DNA is a better marker of a nuclear bleb than lamin B levels that vary widely.

cell biology↗

bioRxiv2023

CTCF is essential for proper mitotic spindle structure and anaphase segregation

Mitosis is an essential process in which the duplicated genome is segregated equally into two daughter cells. CTCF has been reported to be present in mitosis but its importance for mitotic fidelity remains to be determined. To evaluate the importance of CTCF in mitosis, we tracked mitotic behaviors in wild type and two different CTCF CRISPR-based genetic knockdowns. We find that knockdown of CTCF results in prolonged mitoses and failed anaphase segregation via time lapse imaging of SiR-DNA. CTCF knockdown did not alter cell cycling or the mitotic checkpoint, which was activated upon nocodazole treatment. Immunofluorescence imaging of the mitotic spindle in CTCF knockdowns revealed disorganization via tri/tetrapolar spindles and chromosomes behind the spindle pole. Imaging of interphase nuclei showed that nuclear size increased drastically, consistent with failure to divide the duplicated genome in anaphase. Population measurements of nuclear shape in CTCF knockdowns do not display decreased circularity or increased nuclear blebbing relative to wild type. However, failed mitoses do display abnormal nuclear morphologies relative to successful mitoses, suggesting population images do not capture individual behaviors. Thus, CTCF is important for both proper metaphase organization and anaphase segregation which impacts the size and shape of the interphase nucleus.

cell biology↗