[PhD Thesis Presentation_Zoom] - Ms. Sandrine Burriel - "Functional analysis of LMTK1 in lung adenocarcinoma"
Presenter: Ms. Sandrine Burriel
Supervisor: Prof. Tadashi Yamamoto
Co-supervisor: Prof. Nicholas Luscombe
Unit: Cell Signal Unit
Title: Functional analysis of LMTK1 in lung adenocarcinoma
Kinases are master regulators of key cellular processes, including those controlling cell proliferation and the cell cycle. Mutations, abnormal gene expression and aberrant activity of kinases are known to contribute to the development and progression of cancer.
The lemur tail kinase (LMTK) family was discovered 20 years ago but its function in health and disease, particularly in the context of cancer, remains largely unknown. The founding member of the family, LMTK1, also known as apoptosis-associated tyrosine kinase (AATK/AATYK), possesses two splice variants: a transmembrane protein (LMTK1B) and a cytosolic protein (LMTK1A). LMTK1 is predominantly expressed in the brain where it regulates axon and dendrite formation in neurons through its kinase activity. Independent of its function as a kinase, LMTK1 has also been found to act as a scaffold protein, recruiting protein phosphatase 1 (PP1) and SPAK to regulate the activity of the NKCC1 cotransporter. Recent work has established that, in several types of cancers, LMTK1 is downregulated due to DNA methylation of the associated AATK gene, which contributes to cancer progression.
In this project, we investigated the role of LMTK1 lung cancer progression, using a combination of computational analyses based on patient data from The Cancer Genome Atlas and experimental approaches, including RNA sequencing and biochemical analyses. Through patient data analysis, we confirmed that LMTK1 mRNA expression is downregulated in the tumors of non-small cell lung cancer patients, correlating with a moderate impact on survival for lung adenocarcinoma patients. In a lung adenocarcinoma cell model (A549 cells), restoring expression of both splice variants of LMTK1 reduces cell proliferation in a comparable manner, pointing to similar functions for both variants in the context of cell growth. Mutating either the kinase active site or the PP1 docking site showed that, in A549 cells, the kinase activity, rather than the protein scaffold function, is the main driver of reduced proliferation. After hydroxyurea mediated synchronization of cells in late G1 phase, cell cycle analysis showed that LMTK1A expression leads to slower progression from G1 through S phase, concomitant with decreased expression of Cyclin E1 and increased expression of p21. E2F1, a transcription factor known to regulate Cyclin E1 expression and itself target of YAP transcriptional activity, was also found to be decreased. Immunofluorescence showed that wild-type LMTK1A expression dramatically slowed nuclear translocation of YAP, and this effect was abrogated by the kinase-negative LMTK1A mutant. Our data suggest that this effect of LMTK1A might be independent of Rab11A activity, but that LMTK1A may mediate interactions between CD44, Merlin and Akt leading to stabilization of the actin cytoskeleton and enhanced regulation of YAP nuclear translocation. We hypothesise a novel role for LMTK1A kinase activity as a regulator of the Hippo pathway.