To elucidate the effects of neoadjuvant chemotherapy (NAC), we conduct whole transcriptome profiling coupled with histopathology analyses of a longitudinal breast cancer cohort of 146 patients including 110 pairs of serial tumor biopsies collected before treatment, after the first cycle of treatment and at the time of surgery. Here, we show that cytotoxic chemotherapies induce dynamic changes in the tumor immune microenvironment that vary by subtype and pathologic response. Just one cycle of treatment induces an immune stimulatory microenvironment harboring more tumor infiltrating lymphocytes (TILs) and up-regulation of inflammatory signatures predictive of response to anti-PD1 therapies while residual tumors are immune suppressed at end-of-treatment compared to the baseline. Increases in TILs and CD8+ T cell proportions in response to NAC are independently associated with pathologic complete response. Further, on-treatment immune response is more predictive of treatment outcome than immune features in paired baseline samples although these are strongly correlated.

Controlled turnover of proteins as mediated by the ubiquitin proteasome system (UPS) is an important element in plant defense against environmental and pathogen stresses. E3 ligases play a central role in subjecting proteins to hydrolysis by the UPS. Recently, it has been demonstrated that a specific class of E3 ligases termed the U-box ligases are directly associated with the defense mechanisms against abiotic and biotic stresses in several plants. However, no studies on U-box E3 ligases have been performed in one of the important staple crops, barley. In this study, we identified 67 putative U-box E3 ligases from the barley genome and expressed sequence tags (ESTs). Similar to Arabidopsis and rice U-box E3 ligases, most of barley U-box E3 ligases possess evolutionary well-conserved domain organizations. Based on the domain compositions and arrangements, the barley U-box proteins were classified into eight different classes. Along with this new classification, we refined the previously reported classifications of U-box E3 ligase genes in Arabidopsis and rice. Furthermore, we investigated the expression profile of 67 U-box E3 ligase genes in response to drought stress and pathogen infection. We observed that many U-box E3 ligase genes were specifically up-and-down regulated by drought stress or by fungal infection, implying their possible roles of some U-box E3 ligase genes in the stress responses. This study reports the classification of U-box E3 ligases in barley and their expression profiles against drought stress and pathogen infection. Therefore, the classification and expression profiling of barley U-box genes can be used as a platform to functionally define the stress-related E3 ligases in barley.

To develop in vitro respiratory models, it is crucial to identify the factors involved in epithelial cell differentiation. In this study, we comprehensively analyzed the effects of air–liquid interface (ALI) culture on epithelial cell differentiation using single-cell RNA sequencing (scRNA-seq). ALI culture induced a pronounced shift in cell composition, marked by a fivefold increase in ciliated cells and a reduction of more than half in basal cells. Transcriptional signatures associated with epithelial cell differentiation, analyzed using iPathwayGuide software, revealed the downregulation of VEGFA and upregulation of CDKN1A as key signals for epithelial differentiation. Our findings highlight the efficacy of the ALI culture for replicating the human lung airway epithelium and provide valuable insights into the crucial factors that influence human ciliated cell differentiation.

Fibrous dysplasia (FD) is a rare bone disorder characterized by the replacement of normal bone with benign fibro-osseous tissue. Developments in our understanding of the pathophysiology and treatment options are impeded by the lack of suitable research models. In this study, we developed an in vitro organotypic model capable of recapitulating key intrinsic and phenotypic properties of FD. Initially, transcriptomic profiling of individual cells isolated from patient lesional tissues unveiled intralesional molecular and cellular heterogeneity. Leveraging these insights, we established patient-derived organoids (PDOs) using primary cells obtained from patient FD lesions. Evaluation of PDOs demonstrated preservation of fibrosis-associated constituent cell types and transcriptional signatures observed in FD lesions. Additionally, PDOs retained distinct constellations of genomic and metabolic alterations characteristic of FD. Histological evaluation further corroborated the fidelity of PDOs in recapitulating important phenotypic features of FD that underscore their pathophysiological relevance. Our findings represent meaningful progress in the field, as they open up the possibility for in vitro modeling of rare bone lesions in a three-dimensional context and may signify the first step towards creating a personalized platform for research and therapeutic studies.

Small molecule inhibitors of BRAF and MEK have proven effective at inhibiting tumor growth in melanoma patients, however this efficacy is limited due to the almost universal development of drug resistance. To provide advanced insight into the signaling responses that occur following kinase inhibition we have performed quantitative (phospho)-proteomics of human melanoma cells treated with either dabrafenib, a BRAF inhibitor; trametinib, a MEK inhibitor or SCH772984, an ERK inhibitor. Over nine experiments we identified 7827 class I phosphorylation sites on 4960 proteins. This included 54 phosphorylation sites that were significantly down-modulated after exposure to all three inhibitors, 34 of which have not been previously reported. Functional analysis of these novel ERK targets identified roles for them in GTPase activity and regulation, apoptosis and cell-cell adhesion. Comparison of the results presented here with previously reported phosphorylation sites downstream of ERK showed a limited degree of overlap suggesting that ERK signaling responses may be highly cell line and cue specific. In addition we identified 26 phosphorylation sites that were only responsive to dabrafenib. We provide further orthogonal experimental evidence for 3 of these sites in human embryonic kidney cells over-expressing BRAF as well as further computational insights using KinomeXplorer. The validated phosphorylation sites were found to be involved in actin regulation, which has been proposed as a novel mechanism for inhibiting resistance development. These results would suggest that the linearity of the BRAF-MEK-ERK module is at least context dependent.