Although long noncoding RNAs (lncRNAs) predominately reside in nuclear and exert their functions in many biological processes, their potential involvement in cytoplasmic signal transduction remains unexplored. Here, we identified a cytoplasmic lncRNA, Long-Intergenic Noncoding RNA for Kinase Activation (LINK-A), which mediates HB-EGF triggered, EGFR:GPNMB heterodimer-dependent HIF1α phosphorylation at Tyr565 and Ser797 by BRK and LRRK2 respectively. These events cause HIF1α stabilization, HIF1α-p300 interaction, and activation of HIF1α transcriptional programs under normoxic conditions. Mechanistically, LINK-A facilitates the recruitment of BRK to EGFR:GPNMB complex and BRK kinase activation. The BRK-dependent HIF1α Tyr565 phosphorylation interferes with Pro564 hydroxylation, leading to normoxic HIF1α stabilization. Both LINK-A and LINK-A-dependent signaling pathway activation correlate with TNBC, promoting breast cancer glycolysis reprogramming and tumorigenesis. Our findings illustrate the magnitude and diversity of cytoplasmic lncRNAs in signal transduction and highlight the important roles of lncRNAs in cancer.

Background Cisplatin-based neoadjuvant chemotherapy (NAC) before cystectomy is the standard of care for muscle-invasive bladder cancer (MIBC), with 25–50% of patients expected to achieve a pathologic response. Validated biomarkers predictive of response are currently lacking. Objective To discover and validate biomarkers predictive of response to NAC for MIBC. Design, setting, and participants Pretreatment MIBC samples prospectively collected from patients treated in two separate clinical trials of cisplatin-based NAC provided the discovery and validation sets. DNA from pretreatment tumor tissue was sequenced for all coding exons of 287 cancer-related genes and was analyzed for base substitutions, indels, copy number alterations, and selected rearrangements in a Clinical Laboratory Improvements Amendments–certified laboratory. Outcome measurements and statistical analysis The mean number of variants and variant status for each gene were correlated with response. Variant data from the discovery cohort were used to create a classification tree to discriminate responders from nonresponders. The resulting decision rule was then tested in the independent validation set. Results and limitations Patients with a pathologic complete response had more alterations than those with residual tumor in both the discovery (p = 0.024) and validation (p = 0.018) sets. In the discovery set, alteration in one or more of the three DNA repair genes ATM, RB1, and FANCC predicted pathologic response (p < 0.001; 87% sensitivity, 100% specificity) and better overall survival (p = 0.007). This test remained predictive for pathologic response in the validation set (p = 0.033), with a trend towards better overall survival (p = 0.055). These results require further validation in additional sample sets. Conclusions: Genomic alterations in the DNA repair-associated genes ATM, RB1, and FANCC predict response and clinical benefit after cisplatin-based chemotherapy for MIBC. The results suggest that defective DNA repair renders tumors sensitive to cisplatin. Patient summary Chemotherapy given before bladder removal (cystectomy) improves the chance of cure for some but not all patients with muscle-invasive bladder cancer. We found a set of genetic mutations that when present in tumor tissue predict benefit from neoadjuvant chemotherapy, suggesting that testing before chemotherapy may help in selecting patients for whom this approach is recommended.

Phosphatidylinositol-3,4,5-trisphosphate (PIP3) mediates signaling pathways as a second messenger in response to extracellular signals. Although primordial functions of phospholipids and RNAs have been hypothesized in the “RNA world”, physiological RNA-phospholipid interactions and their involvement in essential cellular processes has remained a mystery. We explicate the contribution of lipid-binding long non-coding RNAs (lncRNAs) in cancer cells. Among them, Long Intergenic Noncoding RNA for Kinase Activation (LINK-A) directly interacts with AKT pleckstrin homology domain and PIP3 at the single nucleotide level, facilitating AKT-PIP3 interaction and consequent enzymatic activation. LINK-A-dependent AKT hyperactivation leads to tumorigenesis and resistance to AKT inhibitors. Genomic deletions of the LINK-A PIP3-binding motif dramatically sensitized breast cancer cells to AKT inhibitors. Furthermore, meta-analysis showed the correlation between LINK-A expression and incidence of a SNP (rs12095274: A>G), AKT phosphorylation status, and poor outcomes for breast and lung cancer patients. PIP3-binding lncRNA modulates AKT activation with broad clinical implications.

Bone metastases remain as a serious health concern because of limited therapeutic options. Here, we report that crosstalk between ROR1-HER3 and the Hippo-YAP pathway promotes breast cancer bone metastasis in a long noncoding RNA-dependent fashion. Mechanistically, the orphan receptor tyrosine kinase ROR1 phosphorylates HER3 at a previously unidentified site Tyr1307, upon neuregulin stimulation, independently of other ErbB family members. p-HER3 Tyr1307 recruits the LLGL2-MAYA-NSUN6 RNA-protein complex to methylate Hippo/MST1 at Lys59. This methylation leads to MST1 inactivation and activation of YAP target genes in tumor cells, which elicits osteoclast differentiation and bone metastasis. Furthermore, increased ROR1, p-HER3 Tyr1307 and MAYA levels correlate with tumor metastasis and unfavorable outcomes. Our data provide insights into the mechanistic regulation and linkage of the ROR1-HER3 and Hippo-YAP pathway in cancer-specific context, and also imply valuable therapeutic targets for bone metastasis and possible therapy-resistant tumors.

We describe here three CD19− B cell precursor populations in mouse bone marrow identified using 12-color flow cytometry. Cell transfer experiments indicate lineage potentials consistent with multilineage progenitor (MLP), common lymphoid progenitor (CLP), and B lineage–restricted pre-pro–B (Fr. A), respectively. However, single cell in vitro assays reveal lineage plasticity: lymphoid/myeloid lineage potential for CLP and B/T lineage potential for Fr. A. Despite myeloid potential, recombination activating gene 2 reporter activation is first detected at low levels in most MLP cells, with 95% of CLPs showing 10-fold increased levels. Furthermore, single cell analysis shows that half of CLP and 90% of Fr. A cells contain heavy chain DJ rearrangements. These data, together with expression profiles of lineage-specific genes, demonstrate progressive acquisition of B lineage potential and support an asynchronous view of early B cell development, in which B lineage specification initiates in the MLP/CLP stage, whereas myeloid potential is not lost until the pre-pro–B (Fr. A) stage, and B/T lymphoid plasticity persists until the CD19+ pro–B stage. Thus, MLP, CLP, and Fr. A represent progressively B lineage–specified stages in development, before the CD19+ B lineage–committed pro–B stage.

Zhou et al. demonstrate a requirement for the Let-7–Lin28b axis regulating a shift in development between fetal liver and bone marrow B lymphocyte progenitors in the generation of B1 versus B2 B cells. Specifically, the transcription factor Arid3a, induced by Lin28b and a target of Let-7 miRNA, is sufficient to recapitulate fetal B cell development from bone marrow progenitors.

Intrinsic and acquired cellular resistance factors limit the efficacy of most targeted cancer therapeutics. Synthetic lethal screens in lower eukaryotes suggest that networks of genes closely linked to therapeutic targets would be enriched for determinants of drug resistance. We developed a protein network centered on the epidermal growth factor receptor (EGFR), which is a validated cancer therapeutic target, and used siRNA screening to comparatively probe this network for proteins that regulate the effectiveness of both EGFR-targeted agents and nonspecific cytotoxic agents. We identified subnetworks of proteins influencing resistance, with putative resistance determinants enriched among proteins that interacted with proteins at the core of the network. We found that EGFR antagonists and clinically relevant drugs targeting proteins connected in the EGFR network, such as the kinases protein kinase C or Aurora kinase A, or the transcriptional regulator STAT3, synergized to reduce cell viability and tumor size, suggesting the potential for a direct path to clinical exploitation. Such a focused approach can potentially improve the coherent design of combination cancer therapies.

A common polymorphism at codon 72 in the p53 tumor suppressor gene encodes either proline (P72) or arginine (R72). Several groups have reported that in cultured cells, this polymorphism influences p53's transcriptional, senescence, and apoptotic functions. However, the impact of this polymorphism within the context of a living organism is poorly understood. We generated knock-in mice with the P72 and R72 variants and analyzed the tissues of these mice for apoptosis and transcription. In the thymus, we find that the P72 variant induces increased apoptosis following ionizing radiation, along with increased transactivation of a subset of p53 target genes, which includes murine Caspase 4 (also called Caspase 11), which we show is a direct p53 target gene. Interestingly, the majority of genes in this subset have roles in inflammation, and their promoters contain NF-B binding sites. We show that caspase 4/11 requires both p53 and NF-B for full induction after DNA damage and that the P72 variant shows increased interaction with p65 RelA, a subunit of NF-B. Consistent with this, we show that P72 mice have a markedly enhanced response to inflammatory challenge compared to that of R72 mice. Our data indicate that the codon 72 polymorphism impacts p53's role in inflammation.