Biology subjects

Alfandari, D.

Publications and source records attributed to Alfandari, D..

8 recordsLinked to original sources

bioRxiv2026

Planar polarization of endogenous ADIP during Xenopus neurulation

Coordinated cell polarity and force-responsive protein localization are essential for tissue morphogenesis, yet how embryonic cells sense forces and respond to mechanical cues remains a challenging question. Afadin- and alpha-actinin-binding protein (ADIP) has been implicated in microtubule minus-end anchoring, centrosome maturation and ciliogenesis. ADIP is also proposed to associate with the actomyosin cortex and regulate collective cell migration. ADIP behaves as a mechanosensitive planar cell polarity (PCP) protein when overexpressed in Xenopus embryos, but the distribution and regulation of endogenous ADIP has been unknown. Here we show that ADIP is present in early ectoderm as randomly distributed puncta that rapidly reorganize and polarize during epithelial wound repair. Endogenous ADIP also becomes enriched and planar polarized in the anterior neural plate towards the midline, consistent with its regulation by mechanical forces that operate during neural tube closure. ADIP polarization is attenuated by depletion of the core PCP component Diversin/Ankrd6, in agreement with the proposed interaction between the two proteins during PCP establishment. Finally, pharmacological disruption of microtubules, F-actin, and nonmuscle myosin II eliminates ADIP polarization in the neuroectoderm, indicating roles for microtubules and actomyosin networks in PCP. Together, these findings suggest that endogenous ADIP senses mechanical cues via the cytoskeletal machinery and functions in the context-dependent manner to control collective cell behaviors during vertebrate morphogenesis.

developmental biology↗

bioRxiv2025

Mechanical cues organize planar cell polarity during vertebrate morphogenesis and embryonic wound repair

Planar cell polarity (PCP) is a phenomenon of coordinated cell orientation in many epithelia and is required for early morphogenetic events, such as vertebrate gastrulation or neural tube closure that place embryonic tissues in their proper locations. Known PCP complexes segregate to opposite edges of each cell due to regulatory feedback interactions, however, whether and how PCP is connected to the tension-sensing machinery has been elusive. Here we observed dynamic polarization of Afadin- and -Actinin-interacting protein (ADIP) in the epithelia adjacent to the involuting marginal zone and the folding neural plate of Xenopus embryos, suggesting that it is controlled by mechanical cues. Supporting this hypothesis, ADIP puncta relocated in response to the pulling forces of neighboring ectoderm cells undergoing apical constriction. Moreover, ADIP puncta rapidly polarized in embryos subjected to stretching and during embryonic wound healing. ADIP formed a mechanosensitive complex with the PCP protein Diversin that was distinct from known core PCP complexes and required for wound repair. We propose that mechanically controlled planar polarization of the ADIP-Diversin complex guides cell behaviors in normal morphogenesis and during wound healing. HighlightsO_LIADIP is a mechanosensing protein that is planar polarized together with Diversin in epithelia during morphogenesis and wound repair C_LIO_LIEpithelial wound healing redistributes ADIP-containing complexes towards the wound. C_LIO_LIADIP and Diversin are interdependent and form a complex regulating morphogenesis and wound healing C_LIO_LIPositive regulatory feedback between mechanical forces and PCP signaling drives morphogenesis C_LI

developmental biology↗

bioRxiv2025

Bacterial ubiquitin ligase engineered for small molecule and protein target identification

The Legionella SidE effectors ubiquitinate host proteins independently of the canonical E1-E2 cascade. Here we engineer the SidE ligases to develop a modular proximity ligation approach for the identification of targets of small molecules and proteins, which we call SidBait. We validate the method with known small molecule-protein interactions and use it to identify CaMKII as an off-target interactor of the breast cancer drug ribociclib. Structural analysis and activity assays confirm that ribociclib binds the CaMKII active site and inhibits its activity. We further customize SidBait to identify protein-protein interactions, including substrates for enzymes, and discover the F-actin capping protein (CapZ) as a target of the Legionella effector RavB during infection. Structural and biochemical studies indicate that RavB allosterically binds CapZ and decaps actin, thus functionally mimicking eukaryotic CapZ interacting proteins. Collectively, our results establish SidBait as a reliable tool for identifying targets of small molecules and proteins.

molecular biology↗

bioRxiv2025

A Dapl1+ subpopulation of naive CD8 T cells contains committed precursors of memory lineage.

Memory CD8 T cells play a vital role in providing lasting immune protection, yet their origins remain incompletely understood. Contrary to classical models, emerging evidence suggests that heterogeneity within the naive T cell pool may influence fate decisions prior to antigen encounter. However, the markers of naive T cell heterogeneity have not yet been clearly defined. Here, we describe intraclonal heterogeneity within the naive T cell population marked by the protein Dapl1. Using novel monoclonal antibodies and a reporter-knockout mouse model, we found that Dapl1-positive naive CD8 T cells exhibit distinct phenotypes compared to their Dapl1-negative counterparts. Furthermore, this population includes a subset of pre-programmed precursors biased toward memory lineage fate. The differentiation of these precursors is independent of Dapl1 but relies on the transcription factor Bcl11b, resulting in the generation of Dapl1-positive central memory-like CD8 T cells in response to infection, and stem-like memory cells in response to cancer. Notably, naive Dapl1-positive T cells originate among mature thymocytes and gradually appear in the periphery within several days after birth. Our findings suggest that committed memory precursors in the Dapl1-positive population may represent an alternative pathway for memory CD8 T cell generation, offering new avenues for therapeutic application. ONE-SENTENCE SUMMARYNaive Dapl1 positive CD8 T cells include a pre-programmed subset biased toward differentiation into memory-like T cells.

immunology↗

bioRxiv2024

ADAM interact with large protein complexes to regulate Histone modification, gene expression and splicing.

Cranial neural crest (CNC) cells are key stem cells that contribute to most of the facial structures in vertebrates. ADAM (A Disintegrin And Metalloprotease) proteins are essential for the induction and migration of the CNC. We have shown that Adam13 associates with the transcription factor Arid3a to regulate gene expression. Here we show that Adam13 modulates Histone modifications in the CNC. We show that Arid3a binding to the tfap2 promoter depends on the presence of Adam13. This association promotes the expression of one tfap2 variant expressed in the CNC that uniquely activates the expression of gene critical for CNC migration. We show that both Adam13 and human ADAM9 associate with proteins involved in histone modification and RNA splicing, a function critically affected by the loss of Adam13. We propose that ADAMs may act as extracellular sensors to modulate chromatin availability, leading to changes in gene expression and splicing.

developmental biology↗

bioRxiv2023

ADAM11 a novel regulator of Wnt and BMP4 signaling in neural crest and cancer.

Cranial neural crest (CNC) cells are induced at the border of the neural plate by a combination of FGF, Wnt, and BMP4 signaling. CNC then migrate ventrally and invade ventral structures where they contribute to craniofacial development. Here we show that a non-proteolytic ADAM, Adam11, originally identified as a putative tumor suppressor binds to proteins of the Wnt and BMP4 signaling pathway. Mechanistic studies concerning these non-proteolytic ADAM lack almost entirely. We show that Adam11 positively regulates BMP4 signaling while negatively regulating {beta}-catenin activity. By modulating these pathways, Adam11 controls the timing of neural tube closure and the proliferation and migration of CNC. Using both human tumor data and mouse B16 melanoma cells, we further show that ADAM11 levels similarly correlate with Wnt or BMP4 activation levels. We propose that ADAM11 preserve naive cells by maintaining low Sox3 and Snail/Slug levels through stimulation of BMP4 and repression of Wnt signaling, while loss of ADAM11 results in increased Wnt signaling, increased proliferation and early epithelium to mesenchyme transition.

developmental biology↗

bioRxiv2019

ER transmembrane protein TMTC3 contributes to O-mannosylation of E-cadherin, Cellular Adherence and Embryonic Gastrulation

Protein glycosylation plays essential roles in protein structure, stability and activity such as cell adhesion. The cadherin superfamily of adhesion molecules carry O-linked mannose glycans at conserved sites and it was recently demonstrated that the TMTC1-4 genes contribute to the addition of these O-linked mannoses. Here, biochemical, cell biological and organismal analysis was used to determine that TMTC3 supports the O-mannosylation of E-cadherin, cellular adhesion and embryonic gastrulation. Using genetically engineered cells lacking all four TMTC genes, overexpression of TMTC3 rescued O-linked glycosylation of E-cadherin and cell adherence. The knockdown of the Tmtcs in Xenopus laevis embryos caused a delay in gastrulation that was rescued by the addition of human TMTC3. Mutations in TMTC3 have been linked to neuronal cell migration diseases including Cobblestone lissencephaly. Analysis of TMTC3 mutations associated with Cobblestone lissencephaly found that three of the variants exhibit reduced stability and missence mutations were unable to complement TMTC3 rescue of gastrulation in Xenopus embryo development. Our study demonstrates that TMTC3 regulates O-linked glycosylation and cadherin-mediated adherence, providing insight into its effect on cellular adherence and migration, as well the basis of TMTC3-associated Cobblestone lissencephaly.

cell biology↗

bioRxiv2018

Basolateral localization of MMP14 drives apicobasal polarity change during EMT independently of its catalytic activity

The transmembrane Matrix Metalloproteinase MMP14/MT1-MMP is known to promote cell migration by cleavage of the extracellular matrix. To initiate migration, epithelial cells need to gain mesenchymal attributes. They reduce cell-cell junctions and apicobasal polarity and gain migratory capabilities. This process is named epithelial-mesenchymal transition (EMT). MMP14s implication in EMT is still ill-defined. We used chick neural crest (NC) cells as a model to explore the function of MMP14 in physiological EMT. Our results show that MMP14 is expressed by chick NC cells. However, it is its subcellular localization, rather than its expression, that correlates with EMT. MMP14 is first apical and switches to basolateral domains during EMT. Loss of function and rescue experiments show that MMP14 is involved in EMT independently of its catalytic activity. It lies downstream of pro-EMT genes and upstream of cell polarity. We found that basolateral localization of MMP14 is required and sufficient to induce polarity change in NC cells and neuroepithelial cells, respectively. These effects on polarity occur without impact on cell-cell adhesion or the extracellular matrix. Overall, our data points to a new function of MMP14 in EMT that will need to be further explored in other systems such as cancer cells.

developmental biology↗