Dataset Information

PCNA has specific functions in regulation of metabolism in haematological cells

ABSTRACT: PCNA´s essential roles in DNA replication and repair are well established, while it’s recently discovered non-canonical roles in cellular signalling and regulation of metabolism are less explored. Recent data has indicated that PCNA has a specific non-canonical role in cells of haematological origin. In the present study we have explored this further at the system level by extensive protein-, gene expression, and metabolite profiling combined with integrated pathway analysis and by 13C label experimentation, Here we show that impairing PCNA´s scaffold functions led to massive changes in cellular signalling, but more importantly, a strong reduction in glycolytic metabolites and nucleoside phosphate pools in haematological cancer cell lines was detected, including multiple myeloma cells and acute myelogen leukaemia cells. This was not observed in cell lines from solid tissues such as kidney, bladder and prostate, nor in primary monocytes. Lipopolysaccharide (LPS) stimulated monocytes on the other hand, responded to treatment similarly to the haematological cancer cells, suggesting that cellular stress is an important factor for the response to the APIM-peptide in haematological cells. Integrated Pathway analysis of the metabolite and protein datasets revealed that protein ubiquitination and antigen presentation pathways are affected in haematopoietic cancer cells upon treatment. Focusing on one multiple myeloma cell line we further verified these trends by gene-expression and show that the consequences of reduced central carbon metabolite pools and energy charge is impaired redox and HIF1A regulation. Altogether, our data suggests that PCNA has an important role in regulation of cytoplasmic stress via regulation of central carbon metabolism in haematological cells that can be targeted in cancer treatment.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Mus Musculus (mouse)

TISSUE(S): Embryo, Kidney Epithelial Cell

SUBMITTER: Animesh Sharma

LAB HEAD: Marit Otterlei

PROVIDER: PXD017474 | Pride | 2023-03-10

REPOSITORIES: Pride

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Publications

PCNA regulates primary metabolism by scaffolding metabolic enzymes.

Røst Lisa M LM Ræder Synnøve B SB Olaisen Camilla C Søgaard Caroline K CK Sharma Animesh A Bruheim Per P Otterlei Marit M

Oncogene 20221223 8

The essential roles of proliferating cell nuclear antigen (PCNA) as a scaffold protein in DNA replication and repair are well established, while its cytosolic roles are less explored. Two metabolic enzymes, alpha-enolase (ENO1) and 6-phosphogluconate dehydrogenase (6PGD), both contain PCNA interacting motifs. Mutation of the PCNA interacting motif APIM in ENO1 (F423A) impaired its binding to PCNA and resulted in reduced cellular levels of ENO1 protein, reduced growth rate, reduced glucose consum ...[more]

PMID: 36564470

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Project description:Higher-order chromosomal organization for transcription regulation is poorly understood in eukaryotes. Using genome-wide Chromatin Interaction Analysis with Paired-End-Tag sequencing (ChIA-PET), we mapped long-range chromatin interactions associated with RNA polymerase II in human cells and uncovered widespread promoter-centered intragenic, extragenic, and intergenic interactions. These interactions further aggregated into higher-order clusters, wherein proximal and distal genes were engaged through promoter-promoter interactions. Most genes with promoter-promoter interactions were active and transcribed cooperatively, and some interacting promoters could influence each other implying combinatorial complexity of transcriptional controls. Comparative analyses of different cell lines showed that cell-specific chromatin interactions could provide structural frameworks for cell-specific transcription, and suggested significant enrichment of enhancer-promoter interactions for cell-specific functions. Furthermore, genetically-identified disease-associated noncoding elements were found to be spatially engaged with corresponding genes through long-range interactions. Overall, our study provides insights into transcription regulation by three-dimensional chromatin interactions for both housekeeping and cell-specific genes in human cells. RNA polymerase II (RNAPII) bound chromatin interactions were extracted with Chromatin Interaction Analysis with Paired-End Tag (ChIA-PET) sequencing, in order to study the transcription regulations with RNAPII-associated long-range chromatin interactions. Five cell lines, namely MCF7 (ATCC# HTB-22), K562 (ATCC# CCL-243), HCT116 (ATCC# CCL-247), HeLa (ATCC# CCL-2.2), and NB4 (Roussel and Lanotte, 2001) (provided by Dr. Sherman Weissman, Yale University), were grown under standard culture conditions and harvested at log phase. Harvested cells were cross-linked using 1% formaldehyde followed by neutralization with 0.2M glycine. Chromatin was isolated and subjected to ChIA-PET protocol as described in Fullwood et al (Fullwood et al: An oestrogen-receptor-alpha-bound human chromatin interactome. Nature 2009, 462(7269):58-64). The ChIA-PET sequence reads were processed and analyzed using ChIA-PET Tool (Li et al: ChIA-PET tool for comprehensive chromatin interaction analysis with paired-end tag sequencing. Genome Biol 2010, 11(2):R22).

Dataset Information

PCNA has specific functions in regulation of metabolism in haematological cells

Publications

PCNA regulates primary metabolism by scaffolding metabolic enzymes.

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