Explore the vast range of titles available.

The coordination between membrane trafficking and actomyosin networks is essential to the regulation of cell and tissue shape. Here, we examine Rab protein distributions during Drosophila epithelial tissue remodeling and show that Rab35 is dynamically planar polarized. Rab35 compartments are enriched at contractile interfaces of intercalating cells and provide the first evidence of interfacial monopolarity. When Rab35 function is disrupted, apical area oscillations still occur and contractile steps are observed. However, contractions are followed by reversals and interfaces fail to shorten, demonstrating that Rab35 functions as a ratchet ensuring unidirectional movement. Although actomyosin forces have been thought to drive interface contraction, initiation of Rab35 compartments does not require Myosin II function. However, Rab35 compartments do not terminate and continue to grow into large elongated structures following actomyosin disruption. Finally, Rab35 represents a common contractile cell-shaping mechanism, as mesoderm invagination fails in Rab35 compromised embryos and Rab35 localizes to constricting surfaces. Various stages of tissue morphogenesis involve the contraction of epithelial surfaces. Here, the authors identify the Rab GTPase Rab35 as an essential component of this contractile process, which functions as a membrane ratchet to ensure unidirectional movement of intercalating cells.

Oriented cell intercalation is an essential developmental process that shapes tissue morphologies through the directional insertion of cells between their neighbors. Previous research has focused on properties of cell–cell interfaces, while the function of tricellular vertices has remained unaddressed. Here, we identify a highly novel mechanism in which vertices demonstrate independent sliding behaviors along cell peripheries to produce the topological deformations responsible for intercalation. Through systematic analysis, we find that the motion of vertices connected by contracting interfaces is not physically coupled, but instead possess strong radial coupling. E-cadherin and Myosin II exist in previously unstudied populations at cell vertices and undergo oscillatory cycles of accumulation and dispersion that are coordinated with changes in cell area. Additionally, peak enrichment of vertex E-cadherin/Myosin II coincides with interface length stabilization. Our results suggest a model in which asymmetric radial force balance directs the progressive, ratcheted motion of individual vertices to drive intercalation. Cells need to come together to form tissues of different shapes and sizes. Cells can move about in different ways to shape the tissues. For example, a process called cell intercalation is vital for creating elongated structures like the spinal cord and inner ear. In intercalation, a cell slots itself between neighboring cells to lengthen tissues in one direction. Most of the work to understand cell intercalation has examined the interfaces that form between two neighboring cells. But there are points called vertices where three cells make contact with each other. Vanderleest, Smits et al. have now used microscopy and computational analysis to examine these contact points, known as vertices, in fruit flies. It was thought that vertices that are connected by a single interface coordinate how they move. However, Vanderleest, Smits et al. now show that these connected vertices move independently of each other. Instead, the movements of unconnected vertices on opposite sides of the cell show coordination. Vanderleest, Smits et al. also found that two proteins build up at the vertices in the early stages of intercalation. One of these, called E-cadherin, enables cells to stick to each other. The other protein, called Myosin II, helps E-cadherin to localize to the vertices and also enables cells to contract. These results suggest that the vertices help to guide intercalation and changes in cell shape. Tracking the vertices over time revealed that they slide around the surface of the cells. During this sliding the total length of the interfaces that meet at the vertex remains the same – so as one becomes shorter, neighboring interfaces will become longer. This creates a zipper-like movement of the vertices that tugs the cells into line and suggests a new mechanism by which interconnected cells can change shape. Future work will focus on identifying the molecules that specify these unique vertex behaviors.

A patient with the PSEN1 E280A mutation and homozygous for APOE3 Christchurch ( APOE3Ch ) displayed extreme resistance to Alzheimer’s disease (AD) cognitive decline and tauopathy, despite having a high amyloid burden. To further investigate the differences in biological processes attributed to APOE3Ch , we generated induced pluripotent stem (iPS) cell-derived cerebral organoids from this resistant case and a non-protected control, using CRISPR/Cas9 gene editing to modulate APOE3Ch expression. In the APOE3Ch cerebral organoids, we observed a protective pattern from early tau phosphorylation. ScRNA sequencing revealed regulation of Cadherin and Wnt signaling pathways by APOE3Ch , with immunostaining indicating elevated β-catenin protein levels. Further in vitro reporter assays unexpectedly demonstrated that ApoE3Ch functions as a Wnt3a signaling enhancer. This work uncovered a neomorphic molecular mechanism of protection of ApoE3 Christchurch, which may serve as the foundation for the future development of protected case-inspired therapeutics targeting AD and tauopathies.

Mitotic and cytokinetic processes harness cell machinery to drive chromosomal segregation and the physical separation of dividing cells. Here, we investigate the functional requirements for exocyst complex function during cell division in vivo, and demonstrate a common mechanism that directs anaphase cell elongation and cleavage furrow progression during cell division. We show that onion rings (onr) and funnel cakes (fun) encode the Drosophila homologs of the Exo84 and Sec8 exocyst subunits, respectively. In onr and fun mutant cells, contractile ring proteins are recruited to the equatorial region of dividing spermatocytes. However, cytokinesis is disrupted early in furrow ingression, leading to cytokinesis failure. We use high temporal and spatial resolution confocal imaging with automated computational analysis to quantitatively compare wild-type versus onr and fun mutant cells. These results demonstrate that anaphase cell elongation is grossly disrupted in cells that are compromised in exocyst complex function. Additionally, we observe that the increase in cell surface area in wild type peaks a few minutes into cytokinesis, and that onr and fun mutant cells have a greatly reduced rate of surface area growth specifically during cell division. Analysis by transmission electron microscopy reveals a massive build-up of cytoplasmic astral membrane and loss of normal Golgi architecture in onr and fun spermatocytes, suggesting that exocyst complex is required for proper vesicular trafficking through these compartments. Moreover, recruitment of the small GTPase Rab11 and the PITP Giotto to the cleavage site depends on wild-type function of the exocyst subunits Exo84 and Sec8. Finally, we show that the exocyst subunit Sec5 coimmunoprecipitates with Rab11. Our results are consistent with the exocyst complex mediating an essential, coordinated increase in cell surface area that potentiates anaphase cell elongation and cleavage furrow ingression.

INTRODUCTION We discovered that the APOE3 Christchurch (APOE3Ch) variant may provide resistance to Alzheimer's disease (AD). This resistance may be due to reduced pathological interactions between ApoE3Ch and heparan sulfate proteoglycans (HSPGs). METHODS We developed and characterized the binding, structure, and preclinical efficacy of novel antibodies targeting human ApoE‐HSPG interactions. RESULTS We found that one of these antibodies, called 7C11, preferentially bound ApoE4, a major risk factor for sporadic AD, and disrupts heparin‐ApoE4 interactions. We also determined the crystal structure of a Fab fragment of 7C11 and used computer modeling to predict how it would bind to ApoE. When we tested 7C11 in mouse models, we found that it reduced recombinant ApoE‐induced tau pathology in the retina of MAPT*P301S mice and curbed pTau S396 phosphorylation in brains of systemically treated APOE4 knock‐in mice. Targeting ApoE‐HSPG interactions using 7C11 antibody may be a promising approach to developing new therapies for AD.

INTRODUCTION We described a protected case with familial Alzheimer's disease, homozygous for apolipoprotein E3 (APOE3) Christchurch variant (ApoE3Ch), exhibiting low tau protein levels despite genetic predisposition to the disease due to presenilin (PSEN)1‐E280A. We reported the loss of interaction between ApoE3Ch and heparan sulfate proteoglycans (HSPGs) as a critical protective pathway. Here, we characterized differential interacting partners for both wild‐type and Christchurch variants to identify additional protective mechanisms of ApoE3Ch. METHODS We performed pull‐down of mouse brain lysates using His‐tag‐ApoE3 recombinant proteins and determined interacting partners of ApoE3 via mass‐spectrometry. We then performed in vitro and in vivo assays to validate the top interactors. RESULTS We found enhanced binding of ApoE3Ch to tau and Dickkopf‐1 (Dkk1, a WNT/β‐catenin antagonist) that resulted in reduced tau aggregation in vitro. We demonstrated that ApoE3Ch interacts directly with Dkk1 and tau, reducing tau pathology. These findings supported the hypothesis of novel protective effects of direct ApoE3Ch interactions. Highlights ⁠Apolipoprotein E3 (ApoE3) Christchurch variant (ApoE3Ch) exhibits different protein interaction profiles compared to wild‐type ApoE3, as revealed by proteomic analyses and pull‐down experiments. The ApoE3Ch variant alters the protein's interaction with tau, thus affecting its aggregation in a tau biosensor cell assay and the retina of microtubule‐associated protein tau (MAPT*P301S) transgenic mice. ⁠Gene ontology and pathway analyses indicate that ApoE3Ch interactors are associated with brain‐related disorders and specific upstream regulators, including MAPT, a gene encoding for tau. ⁠Protein–protein interaction studies showed increased binding of ApoE3Ch to Dickkopf1 (Dkk1), a Wnt/β‐catenin pathway antagonist, as compared to ApoE3WT, thus indicating that multiple protective mechanisms are regulated by the ApoE3Ch variant Our study uncovers a novel protective effect of the ApoE3Ch variant against tau pathology, thus proposing new insights into Alzheimer's disease mechanisms and potential therapeutic targets

Oriented cell intercalation is an essential developmental process that shapes tissue morphologies through the directional insertion of cells between their neighbors. Intercalary behaviors in the early Drosophila embryo occur through a remodeling of cell topologies, with cells contracting shared AP interfaces to a single point, followed by newly juxtaposed DV cells constructing horizontally-oriented interfaces between them. Previous research has focused on properties of cell-cell interfaces, and led to a model in which actomyosin networks mediate higher line tensions at AP interfaces to direct contraction. However, the contribution of tricellular vertices to tissue elongation remains unclear. This study shows that cell intercalation uses a novel sliding vertex mechanism that physically couples vertices to radially-oriented forces. Through live imaging and quantitative analysis it was observed that the motion of vertices at contracting interfaces is not coupled, but instead vertices demonstrate strong radial coupling across the area of cells. The vertices of AP junctions show independent sliding behaviors along the cell periphery to produce the topological deformations responsible for intercalation. AP junctions undergo ratcheted length changes that are coordinated with cell area oscillations. These results suggest a model in which oscillations in cell area direct the progressive, ratcheted motion of individual vertices to drive oriented cell intercalation and tissue extension in the Drosophila epithelium. In a second study, analysis of germband extension in 4D revealed that interface contraction and T2 formation can initiate from any point along on the apical-basal axis, including basolateral regions microns away from the apical caps that host major Myosin II populations. Intriguingly, interface contractions transition smoothly into elongations without systematic T2 waiting times and at similar contraction and elongation speeds, suggesting that a common mechanism may underlie both phases of intercalation. This study also showed that the major component of tissue elongation arises from the growth of new interfaces. In a third study, the focus was on the role of membrane trafficking during germband extension. The results of this study showed that Rab35 compartments are enriched at contractile interfaces of intercalating cells. When Rab35 function is disrupted, apical area oscillations still occur and contractile steps are observed. However, contractions are followed by reversals and interfaces fail to shorten, demonstrating that Rab35 functions as a ratchet ensuring unidirectional movement. Finally, Rab35 represents a common contractile cell-shaping mechanism, as mesoderm invagination fails in Rab35 compromised embryos and Rab35 localizes to constricting surfaces. In a fourth and final study, the functional requirements for exocyst complex function during cell division in vivo was investigated, and a common mechanism that directs anaphase cell elongation and cleavage furrow progression during cell division was demonstrated. The results of this study show that onion rings (onr) and funnel cakes (fun) encode the Drosophila homologs of the Exo84 and Sec8 exocyst subunits, respectively. In onr and fun mutant cells, cytokinesis is disrupted early in furrow ingression, leading to cytokinesis failure. Computational analysis was used to quantitatively compare wild-type versus onr and fun mutant cells. The results demonstrate that anaphase cell elongation is grossly disrupted in cells that are compromised in exocyst complex function. Additionally, compared to wild-type, onr and fun mutant cells have a greatly reduced rate of surface area growth specifically during cell division.

Mitotic and cytokinetic processes harness cell machinery to drive chromosomal segregation and the physical separation of dividing cells. Here, we investigate the functional requirements for exocyst complex function during cell division in vivo, and demonstrate a common mechanism that directs anaphase cell elongation and cleavage furrow progression during cell division. We show that onion rings (onr) and funnel cakes (fun) encode the Drosophila homologs of the Exo84 and Sec8 exocyst subunits, respectively. In onr and fun mutant cells, contractile ring proteins are recruited to the equatorial region of dividing spermatocytes. However, cytokinesis is disrupted early in furrow ingression, leading to cytokinesis failure. We use high temporal and spatial resolution confocal imaging with automated computational analysis to quantitatively compare wild-type versus onr and fun mutant cells. These results demonstrate that anaphase cell elongation is grossly disrupted in cells that are compromised in exocyst complex function. Additionally, we observe that the increase in cell surface area in wild type peaks a few minutes into cytokinesis, and that onr and fun mutant cells have a greatly reduced rate of surface area growth specifically during cell division. Analysis by transmission electron microscopy reveals a massive build-up of cytoplasmic astral membrane and loss of normal Golgi architecture in onr and fun spermatocytes, suggesting that exocyst complex is required for proper vesicular trafficking through these compartments. Moreover, recruitment of the small GTPase Rab11 and the PITP Giotto to the cleavage site depends on wild-type function of the exocyst subunits Exo84 and Sec8. Finally, we show that the exocyst subunit Sec5 coimmunoprecipitates with Rab11. Our results are consistent with the exocyst complex mediating an essential, coordinated increase in cell surface area that potentiates anaphase cell elongation and cleavage furrow ingression.

Alzheimer′s disease (AD) is the most common cause of dementia among older adults. APOE3 Christchurch (R136S, APOE3Ch) variant homozygosity was reported in an individual with extreme resistance to autosomal dominant AD due to the PSEN1 E280A mutation. This subject had a delayed clinical age at onset and resistance to tauopathy and neurodegeneration despite extremely high amyloid plaque burden. We established induced pluripotent stem (iPS) cell-derived cerebral organoids from this resistant case and from a non-protected kindred control (with PSEN1 E280A and APOE3/3). We used CRISPR/Cas9 gene editing to successfully remove the APOE3Ch to wild type in iPS cells from the protected case and to introduce the APOE3Ch as homozygote in iPS cells from the non-protected case to examine causality. We found significant reduction of tau phosphorylation (pTau 202/205 and pTau396) in cerebral organoids with the APOE3Ch variant, consistent with the strikingly reduced tau pathology found in the resistant case. We identified Cadherin and Wnt pathways as signaling mechanisms regulated by the APOE3Ch variant through single cell RNA sequencing in cerebral organoids. We also identified elevated β-catenin protein, a regulator of tau phosphorylation, as a candidate mediator of APOE3Ch resistance to tauopathy. Our findings show that APOE3Ch is necessary and sufficient to confer resistance to tauopathy in an experimental ex-vivo model establishing a foundation for the development of novel, protected case-inspired therapeutics for tauopathies, including Alzheimer′s.Competing Interest StatementDrs. Arboleda-Velasquez, Quiroz, and Lopera are listed as inventors on a patent application addressing Christchurch-inspired therapeutics. Dr. Arboleda is a co-founder of EPOCH Biotech, an L.L.C. developing Christchurch-inspired therapeutics. Dr. Quiroz serves as a consultant for Biogen.