Biology subjects

Karjalainen, E.

Publications and source records attributed to Karjalainen, E..

4 recordsLinked to original sources

bioRxiv2025

P4HA3 is abundantly expressed during early embryonic development, but does not catalyze the formation of 4-hydroxyproline in type I collagen

4-Hydroxyproline (4Hyp) is crucially important for the collagen triple helix formation and its thermal stability. It is catalyzed by collagen prolyl 4-hydroxylases (C-P4Hs) that are 2{beta}2 tetramers consisting of two identical catalytic -subunits (encoded by P4HA1, P4HA2 and P4HA3) and two {beta}-subunits that are protein disulfide isomerases (PDI). The functions of isoenzymes I and II (P4HA1 and P4HA2) are widely studied but very little is currently known about the role of P4HA3. Here we show that P4ha1 is generally the most abundant isoform in mouse, whereas P4ha3 expression is tissue and developmental stage specific. P4ha3 expression is highest in the developing skeletal system and peaks at E12.5 together with the other isoforms. Postnatal P4ha3 expression is low and mainly present in calvaria, lung, heart and muscle. The P4ha3 expression virtually disappears at or after 1 week of age in all tissues studied. To evaluate the P4HA3 function in collagen synthesis, we produced cells that lack P4HA3 (P4ha3-/-) and cells that have P4HA3 as the only isoform (P4ha1-/-;P4ha2-/-) by CRISPR/Cas9. C-P4H activity assays indicated no 4Hyp formation in type I collagen by the P4ha1-/-;P4ha2-/- cells, suggesting that the remaining isoform P4HA3 does not participate in type I collagen synthesis. The P4ha3-/- cells secreted intact pepsin-resistant collagen that had a similar 4Hyp content and melting temperature as in WT cells, and the secreted collagen assembled correctly into supramolecular collagen fibrils. In conclusion, our results provide new insights into collagen biosynthesis and suggest that prolyl 4-hydroxylation of type I collagen is mediated by P4HA1 and P4HA2 but not by P4HA3.

biochemistry↗

bioRxiv2025

Sustained epithelial interferon signaling modulates incomplete pathologic response in colorectal cancer

Background & AimsPatients with colorectal cancer have heterogeneous clinical responses to chemotherapy, although clinical guidelines advise little variability in treatment selection based on molecular tumor features. Precision oncology research typically utilizes patient-derived tumor organoids (PDTO) to predict clinical outcomes, but such efforts are often not directed towards identification of molecular factors underlying differential responses to therapy. MethodsBulk RNA-sequencing was performed on treatment-naive PDTOs, and gene expression data was combined to drug sensitivity data to identify transcriptomic features associated with low in vitro sensitivity to chemotherapy. Whole-exome sequencing was performed on primary tumors to infer the somatic mutations of PDTOs and used to identify somatic mutations associated with differential in vitro drug responses. Publicly available gene expression and drug sensitivity data sets were used to validate the results. RNA interference was used for functional validation. ResultsPDTOs with low chemosensitivity had high JAK-STAT pathway activity resulting from high expression of interferon-stimulated genes. Evidence from single-cell RNA-sequencing confirmed chemotherapy-induced expression of interferon-stimulated genes in epithelial cells of cancers with partial response. EPSTI1 knockdown decreased cancer cell viability and sensitized cells to chemotherapy. ConclusionsSustained interferon signaling in epithelial cancer cells contributes to incomplete pathologic response in colorectal cancer. The findings highlight the potential of JAK-STAT inhibition or TRAIL pathway activation to enhance chemotherapy efficacy. Future studies investigating pharmacologic modulation of these pathways in preclinical CRC models are needed to determine their viability as therapeutic targets.

cancer biology↗

bioRxiv2023

Collagen prolyl 4-hydroxylases have sequence specificity towards different X-Pro-Gly triplets

Formation of 4-hydroxyproline (4Hyp) in -X-Pro-Gly- collagen sequences is essential for the thermal stability of collagen molecules. 4Hyp formation is catalyzed by collagen prolyl 4-hydroxylases (C- P4H). Here we identify specific roles for the two main C-P4H isoenzymes by 4Hyp analysis of type I and IV collagens. Loss of C-P4H-I mainly affected prolines preceded by an X-position amino acid with a positively charged or a polar uncharged side chain. In contrast, loss of C-P4H-II affected triplets with a negatively charged glutamate or aspartate in the X-position, and their hydroxylation was found to be important as loss of C-P4H-II alone resulted in reduced collagen melting temperature and altered assembly of collagen fibrils and basement membrane. The C-P4H isoenzyme differences in substrate specificity were explained by selective substrate binding to the active site resulting in differences in Km and Vmax values. In conclusion, this study provides a molecular level explanation for the need of multiple C-P4H isoenzymes to generate collagen molecules capable to assemble into intact extracellular matrix structures.

molecular biology↗

bioRxiv2021

COMPARE, an ultra-fast and robust suite for multiparametric screening, identifies phenotypic drug responses in acute myeloid leukemia

Multiparametric phenotypic screening of cells, for example assessing their responses to small molecules or knockdown/knockout of specific genes, is a powerful approach to understanding cellular systems and identifying potential new therapeutic strategies. However, automated tools for analyzing similarities and differences between a large number of tested conditions have not been readily available. Methods designed for clustering cells cannot identify differences between samples effectively. We introduce O_SCPLOWCOMPAC_SCPLOWRO_SCPLOWEC_SCPLOW for ultra-fast and robust analysis of multiparametric high-throughput screening. Applying a mass-aware gridding algorithm using hypercubes, O_SCPLOWCOMPAC_SCPLOWRO_SCPLOWEC_SCPLOW performs automatic and effective similarity comparison for hundreds to thousands of tests and provides information about the treatment effect. Particularly for screening data, O_SCPLOWCOMPAC_SCPLOWRO_SCPLOWEC_SCPLOW is equipped with modules to remove various sources of bias. Benchmarking tests show that O_SCPLOWCOMPAC_SCPLOWRO_SCPLOWEC_SCPLOW can circumvent batch effects and perform a similarity analysis substantially faster than conventional analysis tools. Applying O_SCPLOWCOMPAC_SCPLOWRO_SCPLOWEC_SCPLOW to high-throughput flow cytometry screening data, we were able to distinguish subtle phenotypic drug responses in a human sample and a genetically engineered mouse model with acute myeloid leukemia (AML). O_SCPLOWCOMPAC_SCPLOWRO_SCPLOWEC_SCPLOW revealed groups of drugs with similar responses even though their mechanisms are distinct from each other. In another screening, O_SCPLOWCOMPAC_SCPLOWRO_SCPLOWEC_SCPLOW effectively circumvented batch effects and grouped samples from AML and myelodysplastic syndrome (MDS) patients using clinical flow cytometry data.

cancer biology↗