TWEAK/Fn14 signalling driven super-enhancer reprogramming promotes pro-metastatic metabolic rewiring in triple-negative breast cancer.

Triple Negative Breast Cancer (TNBC) is the most aggressive breast cancer subtype suffering from limited targeted treatment options. Following recent reports correlating Fibroblast growth factor-inducible 14 (Fn14) receptor overexpression in Estrogen Receptor (ER)-negative breast cancers with metastatic events, we show that Fn14 is specifically overexpressed in TNBC patients and associated with poor survival. We demonstrate that constitutive Fn14 signalling rewires the transcriptomic and epigenomic landscape of TNBC, leading to enhanced tumour growth and metastasis. We further illustrate that such mechanisms activate TNBC-specific super enhancers (SE) to drive the transcriptional activation of cancer dependency genes via chromatin looping. In particular, we uncover the SE-driven upregulation of Nicotinamide phosphoribosyltransferase (NAMPT), which promotes NAD+ and ATP metabolic reprogramming critical for filopodia formation and metastasis. Collectively, our study details the complex mechanistic link between TWEAK/Fn14 signalling and TNBC metastasis, which reveals several vulnerabilities which could be pursued for the targeted treatment of TNBC patients.

Aberrant non-canonical NF-κB signalling reprograms the epigenome landscape to drive oncogenic transcriptomes in multiple myeloma.

In multiple myeloma, abnormal plasma cells establish oncogenic niches within the bone marrow by engaging the NF-κB pathway to nurture their survival while they accumulate pro-proliferative mutations. Under these conditions, many cases eventually develop genetic abnormalities endowing them with constitutive NF-κB activation. Here, we find that sustained NF-κB/p52 levels resulting from such mutations favours the recruitment of enhancers beyond the normal B-cell repertoire. Furthermore, through targeted disruption of p52, we characterise how such enhancers are complicit in the formation of super-enhancers and the establishment of cis-regulatory interactions with myeloma dependencies during constitutive activation of p52. Finally, we functionally validate the pathological impact of these cis-regulatory modules on cell and tumour phenotypes using in vitro and in vivo models, confirming RGS1 as a p52-dependent myeloma driver. We conclude that the divergent epigenomic reprogramming enforced by aberrant non-canonical NF-κB signalling potentiates transcriptional programs beneficial for multiple myeloma progression.

Transcriptional Regulation during Aberrant Activation of NF-κB Signalling in Cancer.

The NF-κB signalling pathway is a major signalling cascade involved in the regulation of inflammation and innate immunity. It is also increasingly recognised as a crucial player in many steps of cancer initiation and progression. The five members of the NF-κB family of transcription factors are activated through two major signalling pathways, the canonical and non-canonical pathways. The canonical NF-κB pathway is prevalently activated in various human malignancies as well as inflammation-related disease conditions. Meanwhile, the significance of non-canonical NF-κB pathway in disease pathogenesis is also increasingly recognized in recent studies. In this review, we discuss the double-edged role of the NF-κB pathway in inflammation and cancer, which depends on the severity and extent of the inflammatory response. We also discuss the intrinsic factors, including selected driver mutations, and extrinsic factors, such as tumour microenvironment and epigenetic modifiers, driving aberrant activation of NF-κB in multiple cancer types. We further provide insights into the importance of the interaction of NF-κB pathway components with various macromolecules to its role in transcriptional regulation in cancer. Finally, we provide a perspective on the potential role of aberrant NF-κB activation in altering the chromatin landscape to support oncogenic development.

Heat stress induced arginylation of HuR promotes alternative polyadenylation of Hsp70.3 by regulating HuR stability and RNA binding.

Arginylation was previously found to promote stabilization of heat shock protein 70.3 (Hsp70.3) mRNA and cell survival in mouse embryonic fibroblasts (MEFs) on exposure to heat stress (HS). In search of a factor responsible for these phenomena, the current study identified human antigen R (HuR) as a direct target of arginylation. HS induced arginylation of HuR affected its stability and RNA binding activity. Arginylated HuR failed to bind Hsp70.3 3' UTR, allowing the recruitment of cleavage stimulating factor 64 (CstF64) in the proximal poly-A-site (PAS), generating transcripts with short 3'UTR. However, HuR from Ate1 knock out (KO) MEFs bound to proximal PAS region with higher affinity, thus excluded CstF64 recruitment. This inhibited the alternative polyadenylation (APA) of Hsp70.3 mRNA and generated the unstable transcripts with long 3'UTR. The inhibition of RNA binding activity of HuR was traced to arginylation-coupled phosphorylation of HuR, by check point kinase 2 (Chk2). Arginylation of HuR occurred at the residue D15 and the arginylation was needed for the phosphorylation. Accumulation of HuR also decreased cell viability upon HS. In conclusion, arginylation dependent modifications of HuR maintained its cellular homeostasis, and promoted APA of Hsp70.3 pre-mRNA, during early HS response.

Comparative analysis of Naja kaouthia venom from North-East India and Bangladesh and its cross reactivity with Indian polyvalent antivenoms.

Naja kaouthia is one of the most prevalent medically important snakes of North East India and Bangladesh responsible for most of the bite cases. In this study, an attempt was made to decipher venom variation of Naja kaouthia venom from North East India and Bangladesh. Using multidimensional methods including reverse phase HPLC, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (1D-PAGE) and two-dimensional gel electrophoresis (2D-PAGE), the quantitative differences in venom composition have been revealed. Moreover, tested in-vitro biochemical and biological activities also exhibited differences which could be due to venom variability. Furthermore, neutralization efficacy of commercially available Indian polyvalent antivenoms (Vins, Bharat Serum, Haffkine) was evaluated and the results displayed significant differences in neutralizing efficacy between the antivenoms. Immunoblotting experiments showed antivenom molecules cross reacted with high molecular mass components while poorly reacted towards low molecular mass proteins. Immuno-depletion study demonstrated that Vins polyvalent antivenom was poor in immunocapturing the venom proteins of both North East Indian and Bangladesh origin Naja kaouthia at the ratio of 1:16 (venom: antivenom).

Arginylation regulates adipogenesis by regulating expression of PPARγ at transcript and protein level.

Protein modification by arginylation regulates protein stability, function and interaction. The loss of arginylation disrupts a diverse set of fundamental cellular processes from proliferation to death. In the current study, role of arginylation in cell differentiation is investigated. Using in vitro preadipocyte differentiation model, it was observed that the inhibition or knockout (KO) of arginyltransferase 1 (ATE1) severely hindered differentiation of preadipocytes into mature adipocytes. Absence of arginylation inhibited expression of two key adipogenic transcription factors PPARγ and C/EBPα, and their downstream adipogenic genes (FABP4, GLUT4, PLN1). Arginylation did not affect the induction of C/EBPß and C/EBPδ, the up-stream regulators of PPARγ gene. However, absence of arginylation affected PPARγ protein expression, independent of its transcript level. The constitutive expression of PPARγ1 protein in Ate1 KO cells as well as ATE1 inhibitor treated wild type cells were dampened due to increased proteasome mediated degradation of PPARγ1 in the absence of arginylation in the cells. Taken together these observations suggested arginylation mediated transcriptional regulation of PPARγ and C/EBPα was downstream of C/EBPß and C/EBPδ, and that the arginylation mediated regulation of PPARγ protein expression may play a role in this process. The inhibition of arginylation in mature adipocytes also reduced expression of lipogenesis genes and decreased fat accumulation in differentiated adipocytes. Thus, the current study shows that arginylation is essential for preadipocyte differentiation and maturation which are thought to be key factors in the maintenance of adipose tissue homeostasis.

Protein arginylation regulates cellular stress response by stabilizing HSP70 and HSP40 transcripts.

ATE1-mediated post-translational addition of arginine to a protein has been shown to regulate activity, interaction, and stability of the protein substrates. Arginylation has been linked to many different stress conditions, namely ER stress, cytosolic misfolded protein stress, and nitrosative stress. However, clear understanding about the effect of arginylation in cellular stress responses is yet to emerge. In this study, we investigated the role of arginylation in heat-stress response. Our findings suggest that Ate1 knock out (KO) cells are more susceptible to heat stress compared with its wild-type counterparts due to the induction of apoptosis in KO cells. Gene expression analysis of inducible heat-shock proteins (HSP70.1, HSP70.3, and HSP40) showed induction of these genes in KO cells early in the heat shock, but were drastically diminished at the later period of heat shock. Further analysis revealed that loss of ATE1 drastically reduced the stability of all three HSP mRNAs. These phenotypes were greatly restored by overexpression of Ate1 in KO cells. Our findings show that arginylation plays a protective role during heat stress by regulating HSP gene expression and mRNA stability.