Biology subjects

Goodall, E.

Publications and source records attributed to Goodall, E..

3 recordsLinked to original sources

bioRxiv2026

Cell-matrix mechanosensing and cellular metabolic demand are linked through SKT and mTORC2

Integrin-based adhesion complexes mediate cell adhesion to the extracellular matrix and enable the cell to interpret and respond to both biochemical and mechanical cues. Such cues can affect a cells metabolic rate; however, how mechanical signals are converted into metabolic rate changes is not clear. We identified an uncharacterised protein; Sickle Tail Protein Homolog (KIAA1217), SKT to be enriched in cell-matrix adhesion complexes in stiff microenvironments. Low SKT expression correlates with an improved prognosis in pancreatic ductal adenocarcinoma (PDAC), suggesting an important role for SKT in extracellular matrix dependent tumour progression. Here, we show that SKT interacts with the mechanistic target of rapamycin complex 2 (mTORC2), a pivotal signalling complex in glucose metabolism, cell growth, and survival. SKT recruits mTORC2 to cell-matrix adhesions in a mechanoresponsive manner. SKT mediated mTORC2 signalling from adhesions is required for maintaining glycolytic flux and control of adhesion dynamics. Our findings show that SKT serves as a rheostat that controls metabolic adaptation of cells to their matrix microenvironment. Collectively, our research provides insights into the molecular mechanisms and interplay between cell adhesion and metabolic signalling in complex and stiff tumour microenvironments. The novel functions identified for SKT in cell-matrix adhesions, mTORC2 signalling and glycolysis unveils a signalling axis between the tumour microenvironment and cellular metabolism that are required for PDAC growth and invasion.

cell biology↗

bioRxiv2025

Cyclin-dependent kinase-like 5 (CDKL5) binds to talin and is anchored at the postsynaptic density via direct interaction with PDZ domains

Cyclin-dependent kinase-like 5 (CDKL5) is a serine/threonine kinase essential for brain development and function. Mutations in the CDKL5 gene cause CDKL5 deficiency disorder (CDD), a severe early-onset epileptic encephalopathy characterised by defects in synapse formation and function. Despite extensive research, the molecular mechanisms by which CDKL5 mutations disrupt synaptic function and lead to epilepsy remain unclear. Here, we report that the major neuronal isoform of CDKL5 contains a C-terminal PDZ domain-binding motif. We demonstrate that this motif mediates interactions with the PDZ domains of PSD-95 and SHANK proteins, facilitating the recruitment of CDKL5 to the postsynaptic density. Disruption of CDKL5s PDZ-binding motif results in its mislocalisation and impaired spine formation. Additionally, we show that CDKL5 directly interacts with the mechanosensitive synaptic scaffold protein talin, via the N-terminal kinase domain of CDKL5 and the R8 rod domain of talin. Our findings establish how CDKL5 is targeted to synapses and suggest that its activity may be spatially regulated through talin-mediated mechanical signalling. We propose that the spatial positioning of the CDKL5 kinase domain might be mechanically-operated and regulated by talin domain unfolding. As talin undergoes structural transitions in its force-dependent binary switch domains, the kinase domain bound to R8 would be moved up and down within the synaptic compartment as a function of the changing talin conformation. These insights enhance our understanding of the pathogenic mechanisms underlying CDKL5 variants with premature stop codons and highlight the need to re-evaluate studies that have used C-terminally tagged or the non-PDZ-binding isoform of CDKL5 to assess its neuronal function. O_FIG O_LINKSMALLFIG WIDTH=188 HEIGHT=200 SRC="FIGDIR/small/672337v1_ufig1.gif" ALT="Figure 1"> View larger version (35K): org.highwire.dtl.DTLVardef@a11d72org.highwire.dtl.DTLVardef@19428b7org.highwire.dtl.DTLVardef@1fd1693org.highwire.dtl.DTLVardef@d5299c_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical AbstractC_FLOATNO Cyclin-dependent kinase-like 5 (CDKL5) is a mechanically regulated enzyme. The C-terminal PDZ-binding motif in CDKL5 anchors CDKL5 at the postsynaptic density (PSD). (A) The large unstructured region serves to tether the CDKL5 kinase domain at the synapse. i) The kinase domain is restricted to the blue semi-circle, giving CDKL5 a zone of activity, with radius the length of the tether (>200 nm). ii) Epilepsy-causing mutations that introduce premature stop codons untether CDKL5. (B) The mechanosensitive synaptic scaffold protein talin is also tethered to the membrane by interaction with integrins and/or amyloid precursor protein (APP). The CDKL5-binding domain, R8 is shown in dark green. The force-dependent binary switch domains in the talin rod region open and close in response to force and change the length of the molecule and the location of the CDKL5-binding site. (C) As talin changes its conformation, the CDKL5-binding site on R8 is moved up and down within the synapse, as a result the kinase activity of CDKL5 is targeted to discrete locations within the synapse. Loss of this localisation or incorrect positioning of the kinase within the synapse would result in loss of synchronisation of the synapse in the context of the circuit it is part of and, we propose, result in re-entry circuits and epilepsy. See also Supplementary Movie 1. C_FIG

neuroscience↗

bioRxiv2017

The essential genome of Escherichia coli K-12

Transposon-Directed Insertion-site Sequencing (TraDIS) is a high-throughput method coupling transposon mutagenesis with short-fragment DNA sequencing. It is commonly used to identify essential genes. Single gene deletion libraries are considered the gold standard for identifying essential genes. Currently, the TraDIS method has not been benchmarked against such libraries and therefore it remains unclear whether the two methodologies are comparable. To address this, a high density transposon library was constructed in Escherichia coli K-12. Essential genes predicted from sequencing of this library were compared to existing essential gene databases. To decrease false positive identification of essential gene candidates, statistical data analysis included corrections for both gene length and genome length. Through this analysis new essential genes and genes previously incorrectly designated as essential were identified. We show that manual analysis of TraDIS data reveals novel features that would not have been detected by statistical analysis alone. Examples include short essential regions within genes, orientation-dependent effects and fine resolution identification of genome and protein features. Recognition of these insertion profiles in transposon mutagenesis datasets will assist genome annotation of less well characterized genomes and provides new insights into bacterial physiology and biochemistry.\n\nIMPORTANCEIncentives to define lists of genes that are essential for bacterial survival include the identification of potential targets for antibacterial drug development, genes required for rapid growth for exploitation in biotechnology, and discovery of new biochemical pathways. To identify essential genes in E. coli, we constructed a very high density transposon mutant library. Initial automated analysis of the resulting data revealed many discrepancies when compared to the literature. We now report more extensive statistical analysis supported by both literature searches and detailed inspection of high density TraDIS sequencing data for each putative essential gene for the model laboratory organism, Escherichia coli. This paper is important because it provides a better understanding of the essential genes of E. coli, reveals the limitations of relying on automated analysis alone and a provides new standard for the analysis of TraDIS data.

microbiology↗