👤 Łukasz Majewski

The cell-intrinsic capacity of neurons to regenerate axons requires widespread coordination of the transcriptome, activation of multiple kinases, and reorganization of the cytoskeleton. Axonal repair is also influenced by extrinsic activating factors, such as neurotrophins. Here, we found that the neurotrophin BDNF amplifies multiple neuron-intrinsic programs to foster axonal regeneration in human iPSC-derived lower motor neurons (i Show less

This study explores the cell fate reprogrammability of H3K27M-mutant pediatric high-grade gliomas (pHGG) using neuronal transdifferentiation as a potential targeted therapy. We treated the BT245 patient-derived glioma cell line with pharmacological combinations targeting neuronal differentiation pathways and performed bulk RNA sequencing to characterize gene expression patterns driving cell fate transitions. Our findings reveal that the drug combinations induce transcriptomic changes consistent with differentiation towards neuronal phenotypes, including the upregulation of synaptic and dendritic signaling genes and the downregulation of malignant signatures. In comparison, astrocytic differentiation media (DM) and H3K27M knockout (KO) promote residual astrocytic phenotypes, suggesting neuronal transdifferentiation as a more effective strategy for mitigating tumor aggressiveness and progression. Differentially expressed genes such as GRIK1, GRIN1, NRXN3, NRXN1, CALB2, SCGN, SLC32A1, SLC1A2, KCNC3, and neurodevelopmental regulators including WNT7A, DLX6, ERBB4, ARX, BCL11B, SEMA3C, and FGFBP3 were identified as key markers regulating the neuron-like lineage transition. This study demonstrates that pHGGs can be phenotypically redirected toward neuronal-like identities through modulating cell fate differentiation programs. These findings advance the concept of 'differentiation therapy' as a promising intervention to reduce phenotypic plasticity and malignancy in pHGG ecosystems. While these are early in vitro findings, the potential ability to steer and control glioma cells toward stable, less malignant fates offers promising translational potential for patient-centered targeted therapies. Show less

DOCK7 (dedicator of cytokinesis 7) is a guanidine nucleotide exchange factor (GEF) for Rac1 GTPase that is involved in neuronal polarity and axon generation as well in Schwann cell differentiation and myelination. Recently, we identified DOCK7 as the binding partner of unconventional myosin VI (MVI) in neuronal-lineage PC12 cells and postulated that this interaction could be important in vivo [Majewski et al. (2012) Biochem Cell Biol., 90:565-574]. Herein, we found that MVI-DOCK7 interaction takes also place in other cell lines and demonstrated that MVI cargo domain via its RRL motif binds to DOCK7 C-terminal M2 and DHR2 domains. In MVI knockdown cells, lower Rac1 activity and a decrease of DOCK7 phosphorylation on Tyr1118 were observed, indicating that MVI could contribute to DOCK7 activity. MVI and DOCK7 co-localization was maintained during NGF-stimulated PC12 cell differentiation and observed also in the outgrowths. Also, during differentiation an increase in phosphorylation of DOCK7 as well as of its downstream effector JNK kinase was detected. Interestingly, overexpression of GFP-tagged MVI cargo domain (GFP-GT) impaired protrusion formation indicating that full length protein is important for this process. Moreover, a transient increase in Rac activity observed at 5min of NGF-stimulated differentiation of PC12 cells (overexpressing either GFP or GFP-MVI) was not detected in cells overexpressing the cargo domain. These data indicate that MVI-DOCK7 interaction could have functional implications in the protrusion outgrowth, and full length MVI seems to be important for delivery and maintenance of DOCK7 along the protrusions, and exerting its GEF activity. Show less

Myosin VI (MVI), the only known myosin that walks towards the minus end of actin filaments, is involved in several processes such as endocytosis, cell migration, and cytokinesis. It may act as a transporting motor or a protein engaged in actin cytoskeleton remodelling via its binding partners, interacting with its C-terminal globular tail domain. By means of pull-down technique and mass spectrometry, we identified Dock7 (dedicator of cytokinesis 7) as a potential novel MVI-binding partner in neurosecretory PC12 cells. Dock7, expressed mainly in neuronal cells, is a guanine nucleotide exchange factor (GEF) for small GTPases, Rac1 and Cdc42, which are the major regulators of actin cytoskeleton. MVI-Dock7 interaction was further confirmed by co-immunoprecipitation of endogenous MVI complexed with Dock7. In addition, MVI and Dock7 colocalized in interphase and dividing cells. We conclude that in PC12 cells MVI-Dock7 complexes may function at different cellular locations during the entire cell cycle. Of note, MVI and Dock7 colocalized in primary culture hippocampal neurons also, predominantly in the outgrowths. We hypothesize that this newly identified interaction between MVI and Dock7 may help explain a mechanism for MVI-dependent regulation of actin cytoskeleton organization. Show less

Tricho-rhino-phalangeal syndrome (TRPS) is characterized by craniofacial and skeletal abnormalities. Three subtypes have been described: TRPS I, caused by mutations in the TRPS1 gene on chromosome 8; TRPS II, a microdeletion syndrome affecting the TRPS1 and EXT1 genes; and TRPS III, a form with severe brachydactyly, due to short metacarpals, and severe short stature, but without exostoses. To investigate whether TRPS III is caused by TRPS1 mutations and to establish a genotype-phenotype correlation in TRPS, we performed extensive mutation analysis and evaluated the height and degree of brachydactyly in patients with TRPS I or TRPS III. We found 35 different mutations in 44 of 51 unrelated patients. The detection rate (86%) indicates that TRPS1 is the major locus for TRPS I and TRPS III. We did not find any mutation in the parents of sporadic patients or in apparently healthy relatives of familial patients, indicating complete penetrance of TRPS1 mutations. Evaluation of skeletal abnormalities of patients with TRPS1 mutations revealed a wide clinical spectrum. The phenotype was variable in unrelated, age- and sex-matched patients with identical mutations, as well as in families. Four of the five missense mutations alter the GATA DNA-binding zinc finger, and six of the seven unrelated patients with these mutations may be classified as having TRPS III. Our data indicate that TRPS III is at the severe end of the TRPS spectrum and that it is most often caused by a specific class of mutations in the TRPS1 gene. Show less