Knowing the Human genome better has allowed major advancements in Personalised Medicine. Nowadays, we can know (if we want) the likelihood to develop a given disease and/or how we will react to different pharmacological treatments. Examples of this include diseases like breast cancer (for diagnosis or estimation of likelihood) and lung cancer (for response to treatment), to name just a few.

That said, our genotype does not have the last word. Research in the last couple of decades has shown the power of other regulatory mechanisms, that may enhance or diminish the effect that our genotype will have on our health. Starting from basic healthy life styles, to other more subtle mechanisms, our genotype defines us, but not completely. Above genetics, we have epigenetics… and everything at the protein level. This post will focus on Post-Translational Modifications (PTM), because, after all, it’s the proteins that are the final effectors of a given response to a treatment or to an environmental stimulus.

TFACT™ DNA-BINDING ELISA KITS » OCT2 TFACT™ DNA-BINDING ELISA by Assay Biotechnology. Source: tebu-bio.

Gene expression is regulated by different mechanisms. One of them is the binding of Transcription Factors (TF) to DNA sequences.

Traditionally, the study of TF-DNA interactions is made by several time-consuming and cumbersome: Electrophoretic Mobility Shift Assays (EMSA), Chromatin Immunoprecipitation, Western blotting, and expression of fused target and reporter genes.

ELISA-based formats now allow to have a more precise TF-DNA interaction study in addition to an ease of use.

Costaining of pAkt-1S473 (red) and PECAM-1 (green) after 30 min flow-adaptation and 5 min ortho- (D) or retrograde flow (E). Image was obtained using Cat. No. 039200-301-J34L for p-Akt staining.

Research nowadays aims at working on models as similar as possible to the real physiological status. This includes the modification of cell culture conditions, For example, one should perform cell culture under “real” oxygen levels (e.g. hypoxia, normoxia, physioxia). For circulating cells, shear stress is a key factor, as cells behave in a different way depending on whether they are cultured under static or dynamic conditions.

RalA and RalB GTPases regulate cell motility, morphology, signaling, vesicular trafficking, and endo/exocytosis. The regulation of these functions is critical for the development and spread of cancer, implicating Ral in oncogenesis and metastasis. Both isoforms are integral for Ras-mediated tumorigenesis, metastasis, and invasion. Despite sharing 82% amino acid sequence identity, effectors, and structural/biochemical properties, RalA and RalB have their own unique functions in oncogenesis due to distinct subcellular localization and differential effector interactions. Ral localization, binding partners, and function are regulated by post-translational modifications (PTMs).

In their recent newsletter, Cytoskeleton Inc. summarize recent findings about the relevance of geranylgeranylation, carboxymethylation, palmitoylation, phosphorylation, and ubiquitination in regulating Ral activity, subcellular localization, effector binding, and ultimately, function.

You can download a copy of this newsletter, or if you have any questions or comments, don’t hesitate to get in contact through the form below.

Kits to measure RalA activation

• G-LISA RalA Activation Assay Biochem Kit (colorimetric format)
• RalA Activation Assay Biochem Kit (bead pull-down format)

If you’d like to get an overview about what’s available in the small G protein field, take a look at this Small GTPase product guide.