Anti-NCK1 monoclonal antibody, clone BG7E8

The majority of mutations identified in patients with amelogenesis imperfecta have been mapped to FAM83H. As FAM83H expression is not limited to the enamel, how FAM83H contributes to amelogenesis is still largely unknown. We previously reported that members of the FAM83 family of proteins interact with and regulate the subcellular distribution of the promiscuous serine-threonine protein kinase CK1 family, through their shared Nterminal DUF1669 domains. FAM83H co-localises with CK1 isoforms to speckle-like structures in both the cytoplasm and nucleus. In this report, we show FAM83H, unlike other FAM83 proteins, interacts and colocalises with NCK1/2 tyrosine kinase adaptor proteins. This interaction is mediated by proline-rich motifs within the C-terminus of FAM83H, specifically interacting with the second and third SH3 domains of NCK1/2. Moreover, FAM83H pathogenic AI mutant proteins, which trigger C-terminal truncations of FAM83H, retain their interactions with CK1 isoforms but lose interaction with NCK1/2. These AI mutant FAM83H proteins acquire a nuclear localisation, and recruit CK1 isoforms to the nucleus where CK1 retains its kinase activity. As understanding the constituents of the FAM83H-localised speckles may hold the key to unravelling potential substrates of FAM83H-associated CK1 substrates, we employed a TurboID-based proximity labelling approach and uncovered several proteins including Iporin and BAG3 as potential constituents of the speckles.

Breast cancer stromal compartment, may influence responsiveness to chemotherapy. Our aim was to detect a stromal cell signature (using a direct approach of microdissected stromal cells) associated with response to neoadjuvant chemotherapy (neoCT) in locally advanced breast cancer (LABC). The tumor samples were collected from 44 patients with LABC (29 estrogen receptor (ER) positive and 15 ER negative) before the start of any treatment. Neoadjuvant chemotherapy consisted of doxorubicin and cyclophosphamide, followed by paclitaxel. Response was defined as downstaging to maximum ypT1a-b/ypN0. The stromal cells, mainly composed of fibroblast and immune cells, were microdissected from fresh frozen tumor samples and gene expression profile was determined using Agilent SurePrint G3 Human Gene Expression microarrays. Expression levels were compared using MeV (MultiExperiment Viewer) software, applying SAM (significance analysis of microarrays). To classify samples according to tumor response, the order of median based on confidence statements (MedOr) was used, and to identify gene sets correlated with the phenotype downstaging, gene set enrichment analysis (GSEA). Nine patients presented disease downstaging. Eleven sequences (FDR 17) were differentially expressed, all of which (except H2AFJ) more expressed in responsive tumors, including PTCHD1 and genes involved in abnormal cytotoxic T cell physiology, TOX, LY75, and SH2D1A. The following four pairs of markers could correctly classify all tumor samples according to response: PTCHD1/PDXDC2P, LOC100506731/NEURL4, SH2D1A/ENST00000478672, and TOX/H2AFJ. Gene sets correlated with tumor downstaging (FDR < 0.01) were mainly involved in immune response or lymphocyte activation, including CD47, LCK, NCK1, CD24, CD3E, ZAP70, FOXP3, and CD74, among others. In locally advanced breast cancer, stromal cells may present specific features of immune response that may be associated with chemotherapy response.