Biology subjects

Rappu, P.

Publications and source records attributed to Rappu, P..

5 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

Jagged1 regulates extracellular matrix deposition and remodeling in triple-negative breast cancer

The extracellular matrix (ECM) in the tumor microenvironment plays an essential role in cancer progression. High expression of Jagged1 correlates with poor patient survival and promotes in vivo tumor growth and invasion in triple-negative breast cancer (TNBC). Using transcriptomics, proteomics, and imaging of cancer cell/fibroblast co-cultures, both in vitro and in vivo, we demonstrate that Jagged1-mediated crosstalk between TNBC cells and fibroblasts enhances myofibroblast activation, collagen deposition, and alignment of ECM fibers. Jagged1 increases transforming growth factor beta (TGF{beta}) activity in fibroblast co-cultures and the Jagged1-induced ECM alignment can be prevented by a TGF{beta} inhibitor. Thus, Jagged1 regulates ECM remodeling upstream of TGF{beta}. Furthermore, higher substrate stiffness increases Jagged1 expression, pointing towards a feed-forward loop between Jagged1, ECM stiffness, and TGF{beta}. With the emergence of safe therapeutics targeting specific Notch ligands and receptors, Jagged1 modulation may offer a path to therapeutically target invasive breast cancer.

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

Embigin deficiency leads to delayed embryonic lung development and high neonatal mortality

Embigin (gp70), a transmembrane glycoprotein, has been shown to regulate hematopoietic stem cell and progenitor cell niche. Still, little is known about its expression and function in other organ systems during development or adulthood. By combining immunofluorescence, RNA sequencing, and in vivo mouse models, we show that embigin is highly expressed during development and in adult lung, kidney, epididymis, skin, and testis. Adult Emb-/- mice have a normal lifespan and fertility without apparent pathologies. In contrast, the Emb-/- embryos are significantly smaller than their WT littermates. Markedly increased mortality of the Emb-/- embryos is seen especially during the neonatal period. Embigin is present in the placenta, but placental morphology and gene expression patterns stay unaltered. At E17.5, Emb-/- mice show defective morphogenesis of the lung, low alkaline phosphatase activity in amniotic fluid, and remarkable activation of genes involved in cell proliferation in the lungs. Thus, lung underdevelopment explains the high neonatal mortality. Our work demonstrates the crucial role of embigin during development, and it paves the way to further characterization of embigin in specific organ systems in development and homeostasis. Summary statementEmbigin is a basigin-group transmembrane glycoprotein. In vivo mouse model shows that embigin is crucial for embryonic lung development and neonatal survival.

developmental biology↗

bioRxiv2019

FAK activity sustains intrinsic and acquired ovarian cancer resistance to platinum chemotherapy

Gene copy number changes, cancer stem cell (CSC) increases, and platinum chemotherapy resistance contribute to poor prognosis in patients with recurrent high grade serous ovarian cancer (HGSOC). CSC phenotypes involving Wnt-{beta}-catenin and aldehyde dehydrogenase activities, platinum resistance, and tumor initiating frequency are here associated with spontaneous genetic gains, including genes encoding KRAS, MYC and FAK, in a new murine model of ovarian cancer (KMF). Noncanonical FAK signaling was sufficient to sustain human and KMF tumorsphere proliferation, CSC survival, and platinum resistance. Increased FAK tyrosine phosphorylation occurred in HGSOC patient tumors surviving neo-adjuvant platinum and paclitaxel chemotherapy and platinum resistant tumorspheres acquired FAK dependence for growth. Importantly, combining a pharmacologic FAK inhibitor with platinum overcame chemoresistance and triggered apoptosis in vitro and in vivo. Knockout, rescue, genomic and transcriptomic analyses collectively identified more than 400 genes regulated along a FAK/{beta}-catenin/Myc axis impacting stemness and DNA repair in HGSOC, with 66 genes gained in a majority of Cancer Genome Atlas samples. Together, these results support combinatorial testing of FAK inhibitors for the treatment of recurrent ovarian cancer.\n\nGraphical Summary\n\nO_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=125 SRC=\"FIGDIR/small/594184_ufig1.gif\" ALT=\"Figure 1\">\nView larger version (52K):\norg.highwire.dtl.DTLVardef@20c737org.highwire.dtl.DTLVardef@1f714faorg.highwire.dtl.DTLVardef@823560org.highwire.dtl.DTLVardef@7e6219_HPS_FORMAT_FIGEXP M_FIG C_FIG Key PointsO_LIHigh grade serous ovarian carcinoma tumors contain PTK2 (FAK) 8q24.3 gains associated with prognostic differences.\nC_LIO_LIKMF, a new murine ovarian cancer model with K-Ras, Myc, and FAK gene gains and intrinsic platinum resistance.\nC_LIO_LIFAK activation in tumors surviving platinum chemotherapy promotes cancer stem cell survival.\nC_LIO_LIFAK facilitates a {beta}-catenin-Myc signaling axis controlling gene expression supporting platinum resistance.\nC_LIO_LIFAK activity is essential for KMF tumor growth and is a targetable cellular adaptation of platinum resistance.\nC_LI

cancer biology↗