DCX

Case Study 1: Abiola Abdulrahman Ayanlaja, 2020

Nuclear translocation of several oncogenic proteins have previously been reported, but neither the translocation of doublecortin (DCX) nor the mechanism involved has been studied. DCX is a neuronal microtubule-associated protein (MAP) that is crucial for adult neurogenesis and neuronal migration and has been associated with poor prognosis in gliomas. The researchers probed DCX expression in different grades of glioma tissues and conventional cells via western blotting. Confocal Immunofluorescence was used to detect DCX expression in the cellular compartments, while subcellular fractionation was probed via western blotting. To probe for DCX functions, stable cells expressing high DCX expression or knockdown were generated using CRISPR-Cas9 viral transfection, while plasmid site-directed mutant constructs were used to validate putative nuclear localization sequence (NLS) predicted via conventional algorithms and comparison with classical NLSs. in-silico modeling was performed to validate DCX interactions with import receptors via the selected putative NLS. DCX undergoes nucleocytoplasmic movement via the RanGTPase signaling pathway with an NLS located on the N-terminus between serine47-tyrosine70. This translocation could be stimulated by MARK's phosphorylation of the serine 47 residue flanking the NLS due to aberrant expression of glial cell line-derived neurotrophic factor (GDNF). High expression and nuclear accumulation of DCX improve invasive glioma abilities in-vitro and in-vivo.

Fig1. DCX protein levels as detected by western blotting in non-neoplastic tissues.

Fig2. Pulse shape analysis of U251MG cells expressing DCX mutants compared with the vector showing lower pulse height in DCXNLS2 mutant cells.

Case Study 2: Maryam Moslehi, 2019

Doublecortin X (DCX) plays essential roles in neuronal development via its regulation of cytoskeleton dynamics. This is mediated through direct interactions between its doublecortin (DC) domains (DC1 and DC2) with microtubules (MTs) and indirect association with actin filaments (F-ACT). While the regulatory role of the DCX C-terminus following DC2 has been established, less is known of the possible contributions made by the DCX N-terminus preceding DC1. Here, we assessed the influence of DCX Ser28 within the DCX N-terminus, on the association of DCX with MTs and F-ACT. The researchers compared the cytoskeletal interactions of the DCX S28E phosphomimetic and DCX S28A phospho-resistant mutants and wild-type DCX. Immunoprecipitation and colocalisation analyses indicated increased association of DCX S28E with F-ACT but decreased interaction with MTs, and conversely enhanced DCX S28A association with MTs but decreased association with F-ACT. To evaluate the impact of DCX mutants on cytoskeletal filaments we performed fluorescence recovery after photobleaching (FRAP) studies on SiR-tubulin and β-actin-mCherry and observed comparable tubulin and actin exchange rates in the presence of DCX WT and DCX S28A. However, they observed faster tubulin exchange rates but slower actin exchange rates in the presence of DCX S28E. Moreover, DCX S28E enhanced the association with the actin-binding protein spinophilin (Spn) suggesting the shift to favour association with both F-ACT and Spn in the presence of DCX S28E.

Fig3. Immunoblotting was performed to detect the endogenous α-tubulin and DCX.

Fig4. The time to reach the half-maximal recovery of fluorescence (t1/2) for SiR-tubulin associated with GFP-DCX proteins.

Fig1. Model of a neuron migrating along radial glia. (Amos Gdalyahu, 2004)

DCX involved in several pathways and played different roles in them. We selected most pathways DCX participated on our site, such as Axon guidance, Developmental Biology, L1CAM interactions, which may be useful for your reference. Also, other proteins which involved in the same pathway with DCX were listed below. Creative BioMart supplied nearly all the proteins listed, you can search them on our site.

DCX has several biochemical functions, for example, calcium-dependent protein serine/threonine kinase activity, calmodulin binding, calmodulin-dependent protein kinase activity. Some of the functions are cooperated with other proteins, some of the functions could acted by DCX itself. We selected most functions DCX had, and list some proteins which have the same functions with DCX. You can find most of the proteins on our site.

DCX has direct interactions with proteins and molecules. Those interactions were detected by several methods such as yeast two hybrid, co-IP, pull-down and so on. We selected proteins and molecules interacted with DCX here. Most of them are supplied by our site. Hope this information will be useful for your research of DCX.

GOLGA2; IKZF1; ZBTB5; TRIM27; TRIM23; MEOX1

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