Asymmetric Division Research Articles

Transplantation Stem Cells: From Definition to Opportunities of Clinical Application

The progressive development of embryology, biology, and hematology in the field of experimental research on the biology of SC contribute to the development of new methods and approaches of cellular and tissue therapy for the treatment of pathological conditions and a number of various diseases. To analyze the information about stem cell (SC) biology, which revealed great possibilities of its use as cellular, gene therapy using both embryonic stem cells (ESC) and SC of an adult organism. The application of ESC requires further study of the clinical possibilities of application in clinical practice, which will directly depend on the limits of tolerance in legal and ethical aspects regarding work with embryonic tissues, the development of an appropriate legislative framework for this field of medicine. The materials for processing were published information sources, publications of scientific research. Used methods of systematic and structural-logical analysis, bibliosemantic. It is promising to carry out research on the feasibility of using auto- and allografts of SC hematopoietic tissue obtained from alternative sources, in particular, umbilical cord blood, embryonic liver, bone marrow in clinical transplantation, the development of new transplantation technologies with the use of non-myeloablative modes of conditioning, transplant purification, the use of hematopoietic factors growth of a new generation, vaccination with dendritic cells, etc. Considering the general aspects of the stem cell (self-renewal, differentiation, plasticity, asymmetric division, niche, stromal support), the possibilities of using embryonic stem cells in regenerative medicine and cell therapy open up. The solution to the problem of SC transplantation gives patients a chance for cure and life extension.

Regulation of epidermal stratification and development by basal keratinocytes.

The epidermis is a stratified squamous epithelium distributed in the outermost layer of the skin and is intimately involved in the formation of a physical barrier to pathogens. Basal keratinocytes possess the properties of stem cells and play an essential role in epidermal development and skin damage recovery. Therefore, understanding the molecular mechanism of how basal keratinocytes participate in epidermal development and stratification is vital for preventing and treating skin lesions. During epidermal morphogenesis, the symmetric division of basal keratinocytes contributes to the extension of skin tissues, while their asymmetric division and migration facilitate epidermal stratification. In this review, we summarize the process of epidermal stratification and illustrate the molecular mechanisms underlying epidermal morphogenesis. Furthermore, we discuss the coordination of multiple signaling pathways and transcription factors in epidermal stratification, together with the roles of cell polarity and cell dynamics during the process.

Initial spindle positioning at the oocyte center protects against incorrect kinetochore-microtubule attachment and aneuploidy in mice.

Spindle positioning within the oocyte must be tightly regulated. In mice, the spindle is predominantly assembled at the oocyte center before its migration toward the cortex to achieve the highly asymmetric division, a characteristic of female meiosis. The significance of the initial central positioning of the spindle is largely unknown. We show that initial spindle positioning at the oocyte center is an insurance mechanism to avoid the premature exposure of the spindle to cortical CDC42 signaling, which perturbs proper kinetochore-microtubule attachments, leading to the formation of aneuploid gametes. These findings contribute to understanding why female gametes are notoriously associated with high rates of aneuploidy, the leading genetic cause of miscarriage and congenital abnormalities.

SOX9 Modulates the Transformation of Gastric Stem Cells Through Biased Symmetric Cell Division

Transformation of stem/progenitor cells has been associated with tumorigenesis in multiple tissues, but stem cells in the stomach have been hard to localize. We therefore aimed to use a combination of several markers to better target oncogenes to gastric stem cells and understand their behavior in the initial stages of gastric tumorigenesis. Mouse models of gastric metaplasia and cancer by targeting stem/progenitor cells were generated and analyzed with techniques including reanalysis of single-cell RNA sequencing and immunostaining. Gastric cancer cell organoids were genetically manipulated with clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) for functional studies. Cell division was determined by bromodeoxyuridine-chasing assay and the assessment of the orientation of the mitotic spindles. Gastric tissues from patients were examined by histopathology and immunostaining. Oncogenic insults lead to expansion of SOX9+ progenitor cells in the mouse stomach. Genetic lineage tracing and organoid culture studies show that SOX9+ gastric epithelial cells overlap with SOX2+ progenitors and include stem cells that can self-renew and differentiate to generate all gastric epithelial cells. Moreover, oncogenic targeting of SOX9+SOX2+ cells leads to invasive gastric cancer in our novel mouse model (Sox2-CreERT;Sox9-loxp(66)-rtTA-T2A-Flpo-IRES-loxp(71);Kras(Frt-STOP-Frt-G12D);P53R172H), which combines Cre-loxp and Flippase-Frt genetic recombination systems. Sox9 deletion impedes the expansion of gastric progenitor cells and blocks neoplasia after Kras activation. Although Sox9 is not required for maintaining tissue homeostasis where asymmetric division predominates, loss of Sox9 in the setting of Kras activation leads to reduced symmetric cell division and effectively attenuates the Kras-dependent expansion of stem/progenitor cells. Similarly, Sox9 deletion in gastric cancer organoids reduces symmetric cell division, organoid number, and organoid size. In patients with gastric cancer, high levels of SOX9 are associated with recurrence and poor prognosis. SOX9 marks gastric stem cells and modulates biased symmetric cell division, which appears to be required for the malignant transformation of gastric stem cells.

Polarized branched Actin modulates cortical mechanics to produce unequal-size daughters during asymmetric division

The control of cell shape during cytokinesis requires a precise regulation of mechanical properties of the cell cortex. Only few studies have addressed the mechanisms underlying the robust production of unequal-sized daughters during asymmetric cell division. Here we report that unequal daughter-cell sizes resulting from asymmetric sensory organ precursor divisions in Drosophila are controlled by the relative amount of cortical branched Actin between the two cell poles. We demonstrate this by mistargeting the machinery for branched Actin dynamics using nanobodies and optogenetics. We can thereby engineer the cell shape with temporal precision and thus the daughter-cell size at different stages of cytokinesis. Most strikingly, inverting cortical Actin asymmetry causes an inversion of daughter-cell sizes. Our findings uncover the physical mechanism by which the sensory organ precursor mother cell controls relative daughter-cell size: polarized cortical Actin modulates the cortical bending rigidity to set the cell surface curvature, stabilize the division and ultimately lead to unequal daughter-cell size.

Towards a mechanistic description of PopZ self-assembly from monomers to micron-sized condensates.

We use a combination of in vitro techniques to characterize the formation of phase separated PopZ condensates. The pole-organizing protein Z (PopZ) phase separates into microdomains at the poles of the bacterium Caulobacter crescentus and modulates asymmetric division by selectively sequestering regulatory reaction pathways. PopZ is conserved across α-proteobacteria and plays a key role in cytosol organization. Using mass-photometry, we found that PopZ self-assembles into a distinct distribution of highly stable oligomers in a concentration-dependent manner. Strikingly, PopZ forms trimers and hexamers but seems to only grow beyond hexamers by the addition of more hexamers. Mutations in its oligomerization domain affect the oligomeric distribution as well as the fluidity and saturation concentration of the resulting condensate. PopZ phase separation requires divalent cations, even above its saturation concentration. This allows us to study PopZ at identical concentrations before and after condensation. To characterize the identity of the relevant oligomers and the interparticle interactions that drive phase separation, we study PopZ across three length scales. On the 0.1-1 nanomolar scale, we employ single-molecule FRET to monitor conformational changes within the structured OD and between the OD and the adjacent IDR upon oligomerization and condensation. On the 1-100 nanomolar scale, we use small angle x-ray scattering (SAXS) to monitor the size and shape of PopZ oligomers and microfilaments. Finally, on the micron scale, we study condensates' size, internal and exchange dynamics, and material properties using differential interference contrast and fluorescence microscopies. Combined, our work provides a mechanistic description of PopZ assembly from monomers to micron-sized condensates.

VSELs and OSCs together sustain oogenesis in adult ovaries and their dysfunction results in age-related senescence, PCOS, POI and cancer

Multiple studies using single-cell RNA sequencing (scRNAseq) have failed to detect stem cells in adult ovaries. We have maintained that two populations of ovarian stem cells including pluripotent, very small embryonic-like stem cells (VSELs) and tissue-committed ‘progenitors’ termed ovarian stem cells (OSCs) can easily be detected in Hematoxylin and Eosin-stained ovary surface epithelial (OSE) cells smears prepared from both mice and human ovaries. Most likely the stem cells never get subjected to scRNAseq since they pellet down only by centrifuging cells suspension at 1000 g while cells for scRNAseq were invariably prepared by centrifuging at 200-400 g. A recent article provided further explanation for the failure of scRNAseq to detect ovarian stem cells. Extensive reanalysis of data (generated by scRNAseq) using an advanced software successfully detected OSCs and meiotic markers supporting neo-oogenesis in adult human ovaries. But this article remained critical on the biological relevance of VSELs and their relationship with OSCs. By carefully studying the OSE cells smears (which hold VSELs, OSCs and germ cell nests GCNs), prepared by partial trypsin digestion of intact mice ovaries during different stages of estrus cycle, we have successfully delineated novel functions of VSELs/OSCs in vivo under physiological conditions. VSELs undergo asymmetrical divisions to self-renew and give rise to slightly bigger OSCs which in turn undergo symmetrical divisions and clonal expansion to form GCNs, regular neo-oogenesis and follicle assembly. GCNs have been earlier described in fetal ovaries and during OSE cells culture (from adult ovaries) in response to FSH treatment. Dysfunction of VSELs/OSCs (which express ERα, ERβ, FSHR) due to neonatal exposure to endocrine disruption results in ovarian insufficiency and polycystic ovaries. VSELs have also been implicated in ovarian cancer. Age-related ovarian senescence/menopause is also due to dysfunction and blocked differentiation of VSELs/OSCs. These novel findings in vivo along with abundant in vitro and lineage tracing studies data in published literature provides huge scope for further research, offers novel avenues to manage ovarian pathologies and calls for re-writing of textbooks.

Antibody Uptake Assay for Tracking Notch/Delta Endocytosis During the Asymmetric Division of Zebrafish Radial Glia Progenitors.

Asymmetric cell division (ACD), which produces two daughter cells of different fates, is fundamental for generating cellular diversity. In the developing organs of both invertebrates and vertebrates, asymmetrically dividing progenitors generates a Notchhi self-renewing and a Notchlo differentiating daughter. In the embryonic zebrafish brain, radial glia progenitors (RGPs)-the principal vertebrate neural stem cells-mostly undergo ACD to give birth to one RGP and one differentiating neuron. The optical clarity and easy accessibility of zebrafish embryos make them ideal for in vivo time-lapse imaging to directly visualize how and when the asymmetry of Notch signaling is established during ACD. Recent studies have shown that dynamic endocytosis of the Notch ligand DeltaD plays a crucial role in cell fate determination during ACD, and the process is regulated by the evolutionarily conserved polarity regulator Par-3 (also known as Pard3) and the dynein motor complex. To visualize the in vivo trafficking patterns of Notch signaling endosomes in mitotic RGPs, we have developed this antibody uptake assay. Using the assay, we have uncovered the dynamicity of DeltaD-containing endosomes during RGP division.

Bacterial filaments recover by successive and accelerated asymmetric divisions that allow rapid post-stress cell proliferation.

Filamentation is a reversible morphological change triggered in response to various stresses that bacteria might encounter in the environment, during host infection or antibiotic treatments. Here we re-visit the dynamics of filament formation and recovery using a consistent framework based on live-cells microscopy. We compare the fate of filamentous Escherichia coli induced by cephalexin that inhibits cell division or by UV-induced DNA-damage that additionally perturbs chromosome segregation. We show that both filament types recover by successive and accelerated rounds of divisions that preferentially occur at the filaments' tip, thus resulting in the rapid production of multiple daughter cells with tightly regulated size. The DNA content, viability and further division of the daughter cells essentially depends on the coordination between chromosome segregation and division within the mother filament. Septum positioning at the filaments' tip depends on the Min system, while the nucleoid occlusion protein SlmA regulates the timing of division to prevent septum closure on unsegregated chromosomes. Our results not only recapitulate earlier conclusions but provide a higher level of detail regarding filaments division and the fate of the daughter cells. Together with previous reports, this work uncovers how filamentation recovery allows for a rapid cell proliferation after stress treatment.

Cell Cycle Dynamics during Stomatal Development: Window of MUTE Action and Ramification of Its Loss-of-Function on an Uncommitted Precursor.

Plants develop in the absence of cell migration. As such, cell division and differentiation need to be coordinated for functional tissue formation. Cellular valves on the plant epidermis, stomata, are generated through a stereotypical sequence of cell division and differentiation events. In Arabidopsis, three master regulatory transcription factors, SPEECHLESS (SPCH), MUTE and FAMA, sequentially drive initiation, proliferation and differentiation of stomata. Among them, MUTE switches the cell cycle mode from proliferative asymmetric division to terminal symmetric division and orchestrates the execution of the single symmetric division event. However, it remains unclear to what extent MUTE regulates the expression of cell cycle genes through the symmetric division and whether MUTE accumulation itself is gated by the cell cycle. Here, we show that MUTE directly upregulates the expression of cell cycle components throughout the terminal cell cycle phases of a stomatal precursor, not only core cell cycle engines but also check-point regulators. Time-lapse live imaging using the multicolor Plant Cell Cycle Indicator revealed that MUTE accumulates up to the early G2 phase, whereas its successor and direct target, FAMA, accumulate at late G2 through terminal mitosis. In the absence of MUTE, meristemoids fail to differentiate and their G1 phase elongates as they reiterate asymmetric divisions. Together, our work provides the framework of cell cycle and master regulatory transcription factors to coordinate a single symmetric cell division and suggests a mechanism for the eventual cell cycle arrest of an uncommitted stem-cell-like precursor at the G1 phase.

Cdc42 Regulates the Expression of Cytoskeleton and Microtubule Network Proteins to Promote Invasion and Metastasis of Progeny Cells Derived from CoCl2-induced Polyploid Giant Cancer Cells.

Purpose: Our previous studies have shown that CoCl2 can induce the formation of polyploid giant cancer cells (PGCCs) and PGCCs could produce progeny cells via asymmetric division. In this study, the molecular mechanism by which PGCCs generate progeny cells with high invasion and migration abilities was explored. Methods: In this study, PGCCs induced by CoCl2 produced progeny cells via asymmetric division, which was observed dynamically using laser scanning confocal microscopy. Cell cycle in LoVo and Hct116 before and after CoCl2 treatment was analyzed by flow cytometry. Cell function experiments, co-immunoprecipitation, mass spectrometry analysis, ML141 treatment, western blotting, and siRNA transfection experiments were used to demonstrate that Cdc42/PAK1 was involved in the regulation of cytoskeleton expression. The proliferation, migration, and invasion abilities of PGCCs and progeny cells were compared in PGCCs and progeny cells with and without inhibiting the expression of Cdc42 and PAK1. Results: G2/M phase arrest appeared in CoCl2-treated LoVo and Hct116 cells. After CoCl2 treatment, an increased expression of Cdc42 and PAK1 led to a decrease in the expression of stathmin and an increase in the expression of phosphorylated stathmin, which is located in the nucleus of PGCCs and progeny cells. PTPN14 negatively regulates the expression of PAK1 and p38MAPK. Low levels of PTPN14 expression, a downstream regulatory protein of stathmin, endows progeny tumor cells generated by PGCCs with the ability to invade and metastasize. The expression of PKA1α, cathepsin B, and D increased in CoCl2-treated cells compared with that in the control cells, associated with the infiltration and migration of PGCCs with their progeny cells. Conclusion: CoCl2-induced overexpression of Cdc42 plays a critical role in increasing the infiltration and migration abilities of PGCCs and progeny cells by regulating cytoskeleton protein expression.

Microtubule-mitochondrial attachment facilitates cell division symmetry and mitochondrial partitioning in fission yeast.

Association with microtubules inhibits the fission of mitochondria in Schizosaccharomyces pombe. Here, we show that this attachment of mitochondria to microtubules is an important cell-intrinsic factor in determining cell division symmetry. By comparing mutant cells that exhibited enhanced attachment and no attachment of mitochondria to microtubules (Dnm1Δ and Mmb1Δ, respectively), we show that microtubules in these mutants displayed aberrant dynamics compared to wild-type cells, which resulted in errors in nuclear positioning. This translated to cell division asymmetry in a significant proportion of both Dnm1Δ and Mmb1Δ cells. Asymmetric division in Dnm1Δ and Mmb1Δ cells resulted in unequal distribution of mitochondria, with the daughter cell that received more mitochondria growing faster than the other daughter cell. Taken together, we show the existence of homeostatic feedback controls between mitochondria and microtubules in fission yeast, which directly influence mitochondrial partitioning and, thereby, cell growth. This article has an associated First Person interview with the first author of the paper.

Reduction, proliferation, and differentiation defects of stem cells over time: a consequence of selective accumulation of old centrioles in the stem cells?

All molecules, structures, cells in organisms are subjected to destruction during the process of vital activities. In the organisms of most multicellular animals and humans, the regeneration process always takes place: destruction of old cells and their replacement with the new. The replacement of cells happens even if the cells are in perfect condition. The sooner the organism destroys the cells that emerged a certain time ago and replaces them with the new (i.e., the higher is the regeneration tempo), the younger the organism is. Stem cells are progenitor cells of the substituting young cells. Asymmetric division of a mother stem cell gives rise to one, analogous to the mother, daughter cell, and to a second daughter cell that takes the path of further differentiation. Despite such asymmetric divisions, the pool of stem cells diminishes in its quantity over time. Moreover, intervals between stem cell divisions increase. The combination of these two processes causes the decline of regeneration tempo and aging of the organism. During asymmetric stem cell divisions daughter cells, with preserved potency of the stem cell, selectively conserve mother (old) centrioles. In contrast with molecules of nuclear DNA, reparations do not take place in centrioles. Hypothetically, old centrioles are more subjected to destruction than other structures of a cell-which makes centrioles potentially the main structure of aging.

Opposite polarity programs regulate asymmetric subsidiary cell divisions in grasses.

Grass stomata recruit lateral subsidiary cells (SCs), which are key to the unique stomatal morphology and the efficient plant-atmosphere gas exchange in grasses. Subsidiary mother cells (SMCs) strongly polarise before an asymmetric division forms a SC. Yet apart from a proximal polarity module that includes PANGLOSS1 (PAN1) and guides nuclear migration, little is known regarding the developmental processes that form SCs. Here, we used comparative transcriptomics of developing wild-type and SC-less bdmute leaves in the genetic model grass Brachypodium distachyon to identify novel factors involved in SC formation. This approach revealed BdPOLAR, which forms a novel, distal polarity domain in SMCs that is opposite to the proximal PAN1 domain. Both polarity domains are required for the formative SC division yet exhibit various roles in guiding pre-mitotic nuclear migration and SMC division plane orientation, respectively. Nonetheless, the domains are linked as the proximal domain controls polarisation of the distal domain. In summary, we identified two opposing polarity domains that coordinate the SC division, a process crucial for grass stomatal physiology.

Skeletal Muscle Stem Cells in Aging: Asymmetric/Symmetric Division Switching

In aged muscle, satellite cells’ symmetric and asymmetric divisions are impaired, and intrinsic and extrinsic complex mechanisms govern these processes. This review presents many updated aspects regarding muscle stem cells’ fate in normal and aging conditions. The balance between self-renewal and commitment divisions contributes to muscle regeneration, muscle homeostasis, aging, and disease. Stimulating muscle regeneration in aging could be a therapeutic target, but there is still a need to understand the many mechanisms that influence each other in satellite cells and their niche. We highlight here the general outlines regarding satellite cell divisions, the primary markers present in muscle stem cells, the aging aspects concerning signaling pathways involved in symmetric/asymmetric divisions, the regenerative capacity of satellite cells and their niche alteration in senescent muscle, genetics and epigenetics mechanisms implied in satellite cells aging and exercise effect on muscle regeneration in the elderly.

Maternal mRNA deadenylation and allocation via Rbm14 condensates facilitate vertebrate blastula development.

Early embryonic development depends on proper utilization and clearance of maternal transcriptomes. How these processes are spatiotemporally regulated remains unclear. Here we show that nuclear RNA-binding protein Rbm14 and maternal mRNAs co-phase separate into cytoplasmic condensates to facilitate vertebrate blastula-to-gastrula development. In zebrafish, Rbm14 condensates were highly abundant in blastomeres and markedly reduced after prominent activation of zygotic transcription. They concentrated at spindle poles by associating with centrosomal γ-tubulin puncta and displayed mainly asymmetric divisions with a global symmetry across embryonic midline in 8- and 16-cell embryos. Their formation was dose-dependently stimulated by m6 A, but repressed by m5 C modification of the maternal mRNA. Furthermore, deadenylase Parn co-phase separated with these condensates, and this was required for deadenylation of the mRNAs in early blastomeres. Depletion of Rbm14 impaired embryonic cell differentiations and full activations of the zygotic genome in both zebrafish and mouse and resulted in developmental arrest at the blastula stage. Our results suggest that cytoplasmic Rbm14 condensate formation regulates early embryogenesis by facilitating deadenylation, protection, and mitotic allocation of m6 A-modified maternal mRNAs, and by releasing the poly(A)-less transcripts upon regulated disassembly to allow their re-polyadenylation and translation or clearance.

A Study on the Effects of Gendered Social Norms on the Tradeoff Between Paid and Unpaid Work in Korea.

In this study, Korean time-use survey data for coupled households is analyzed to show that unpaid work time is endogenous in its relationship with paid work time because the views of traditional gender roles affect gender disparity in unpaid work time. The data not only includes time allocation between husbands and wives but also their views of traditional gender roles within their households, and this information can represent gendered social norms that can potentially explain the distribution of unpaid work between husbands and wives. The control function model is estimated to identify the tradeoff between unpaid work time and paid work time by solving the endogeneity problem. The results of this study show that wives' unpaid work is likely to be affected by gendered social norms and that the effect can be larger for those having children. In addition, only in the case of wives, unpaid work time is found to be negatively associated with whether to work full-time, showing that wives' burden of unpaid work could prevent them from working full time. The results indicate that it is crucial to recognize the need to change gendered social norms to address an asymmetric division of unpaid work between husbands and wives.

SMEK1 promotes lung adenocarcinoma proliferation and invasion by activating Wnt/β-catenin signaling pathway.

SMEK1, also known as PP4R3α, the regulatory subunit 3α of serine and threonine phosphatase PP4, participates in diversely critical biological processes such as the integration of centromere, deacetylation of histones, asymmetric divisions of neuroblast, and other crucial cellular activities. SMEK1 was formerly reported to play a part in carcinogenesis. This study aims to reveal the role of SMEK1 in lung adenocarcinoma and the underlying molecular mechanism. Using immunohistochemical (IHC) staining, the protein level of SMEK1 in lung adenocarcinoma and adjacent non-tumor tissue was detected. The functional role of SMEK1 in cell proliferation and invasion was explored using cell counting kit-8 and Transwell assay, respectively. Xenograft tumor experiment was used to investigate the effect of SMEK1 on tumor growth in vivo. The alteration of Wnt/β-catenin signaling pathway was detected by Western blotting, quantitative PCR, and dual-luciferase reporter assays. SMEK1 was highly expressed at the protein level in lung adenocarcinoma compared to the adjacent non-tumor tissue. In vitro, suppression of SMEK1 significantly decreased the proliferation, migration, and invasion of lung adenocarcinoma cell lines, while overexpression of SMEK1 enhanced above abilities. The xenograft model demonstrated that down-regulation of SMEK1 significantly inhibited tumor growth in vivo. In addition, we found that SMEK1 could positively regulate Wnt/β-catenin signaling in lung adenocarcinoma cell lines. SMEK1 exerts a cancer-promoting effect in lung adenocarcinoma by activating Wnt/β-catenin signaling.

Abstract B021: Claudin-1 expression promotes chemoresistance and cancer stem cell characteristics in colorectal cancer

Abstract Background: Claudins are integral components of tight junction complexes. Beyond just passive cell adhesion molecules, they influence structural dynamics, molecular trafficking, and important cell signaling pathways. Claudins are often dysregulated in cancer, leading to changes in adhesion and metastatic potential. Claudin-1 (Cldn1) is expressed at low levels in normal colon but is highly expressed in colorectal cancer (CRC) and promotes epithelial-mesenchymal transition and metastasis. Cancer stem cells (CSCs) are a small subset of cancer cells that display increased capacity for self-renewal and can undergo symmetric and asymmetric division to reform tumors after treatment. CSCs are resistant to cytotoxic drugs and are thought to be responsible for treatment failure, metastasis, and relapse. Significance of problem: CRC is the second leading cause of cancer-related deaths in the United States with an estimated 151,030 new cases in 2022. Although CRC has an overall relative 5-year survival rate of 64%, prognosis varies dramatically with the stage at diagnosis. Localized CRC can be treated surgically and has a 5-year survival rate of 91%, while the survival rate of metastatic CRC falls to 15%. The primary problem in treating metastatic CRC is a combination of intrinsic and acquired chemoresistance. Therefore, understanding the mechanisms of chemoresistance is vital to developing effective therapies. Hypothesis: Claudin-1 expression in colorectal cancer increases resistance to chemotherapeutic drugs and promotes cancer stem cell characteristics. Experimental design: Bioinformatic analysis was performed on a database of responders and non-responders to standard-of-care CRC chemotherapy. Cldn1 expression was manipulated in CRC cell lines, and assays were performed to evaluate chemoresistance and CSC characteristics. Results: Bioinformatic analysis of a patient dataset of responders and non-responders to chemotherapy revealed that Cldn1 expression was positively correlated with resistance to FOLFOX treatment. It was also significantly correlated with colorectal CSC marker expression including CD44, CD133, and Lgr5. Overexpression of Cldn1 in SW480 cells led to a significant increase in resistance to 5-FU, side population and ALDH-high percentage, colorectal CSC markers, and tumorigenic potential, while knockout of Cldn1 in SW620 and DLD-1 cells led to a decreased response in these assays. Follow-up experiments identified the PDZ-binding domain of Cldn1 as essential for the chemoresistant phenotype. Specifically, both CD44 and its downstream target phospho-AKT are increased with the expression of full-length but not mutated Cldn1. Conclusion: Cldn1 expression induces chemoresistance and cancer stem cell properties in colorectal cancer. Because of this, Cldn1 may serve as a biomarker in CRC for prescribing more aggressive adjuvant therapy or as a molecule for targeted treatment aimed at the resistant tumor cells responsible for treatment failure and recurrence. Citation Format: Mark Primeaux, Saiprasad Gowrikumar, Dhundy Bastola, Punita Dhawan, Amar Singh. Claudin-1 expression promotes chemoresistance and cancer stem cell characteristics in colorectal cancer [abstract]. In: Proceedings of the AACR Special Conference on Colorectal Cancer; 2022 Oct 1-4; Portland, OR. Philadelphia (PA): AACR; Cancer Res 2022;82(23 Suppl_1):Abstract nr B021.

Rassf7a promotes spinal cord regeneration and controls spindle orientation in neural progenitor cells.

Spinal cord injury (SCI) can cause long-lasting disability in mammals due to the lack of axonal regrowth together with the inability to reinitiate spinal neurogenesis at the injury site. Deciphering the mechanisms that regulate the proliferation and differentiation of neural progenitor cells is critical for understanding spinal neurogenesis after injury. Compared with mammals, zebrafish show a remarkable capability of spinal cord regeneration. Here, we show that Rassf7a, a member of the Ras-association domain family, promotes spinal cord regeneration after injury. Zebrafish larvae harboring a rassf7a mutation show spinal cord regeneration and spinal neurogenesis defects. Live imaging shows abnormal asymmetric neurogenic divisions and spindle orientation defects in mutant neural progenitor cells. In line with this, the expression of rassf7a is enriched in neural progenitor cells. Subcellular analysis shows that Rassf7a localizes to the centrosome and is essential for cell cycle progression. Our data indicate a role for Rassf7a in modulating spindle orientation and the proliferation of neural progenitor cells after spinal cord injury.