Polarized and Stage-Dependent Distribution of Immunoreactivity for Novel PDZ-Binding Protein Preso1 in Adult Neurogenic Regions

이은수,김운령,김영화,이현우,김현,선웅 대한내분비학회 2014 Endocrinology and metabolism Vol.29 No.3

Background: Adult neural stem cells have the potential for self-renewal and differentiation into multiple cell lineages via symmetric or asymmetric cell division. Preso1 is a recently identified protein involved in the formation of dendritic spines and the promotion of axonal growth in developing neurons. Preso1 can also bind to cell polarity proteins, suggesting a potential role for Preso1 in asymmetric cell division. Methods: To investigate the distribution of Preso1, we performed immunohistochemistry with adult mouse brain slice. Also, polarized distribution of Preso1 was assessed by immunocytochemistry in cultured neural stem cells. Results: Immunoreactivity for Preso1 (Preso1-IR) was strong in the rostral migratory stream and subventricular zone, where proliferating transit-amplifying cells and neuroblasts are prevalent. In cultured neural stem cells, Preso1-IR was unequally distributed in the cell cytosol. We also observed the distribution of Preso1 in the subgranular zone of the hippocampal dentate gyrus, another neurogenic region in the adult brain. Interestingly, Preso1-IR was transiently observed in the nuclei of doublecortin-expressing neuroblasts immediately after asymmetric cell division. Conclusion: Our study demonstrated that Preso1 is asymmetrically distributed in the cytosol and nuclei of neural stem/progenitor cells in the adult brain, and may play a significant role in cell differentiation via association with cell polarity machinery.

Substrate curvature affects the shape, orientation, and polarization of renal epithelial cells

Yu, Sun-Min,Oh, Jung Min,Lee, Junwon,Lee-Kwon, Whaseon,Jung, Woonggyu,Amblard, Franç,ois,Granick, Steve,Cho, Yoon-Kyoung Elsevier 2018 ACTA BIOMATERIALIA Vol.77 No.-

<P><B>Abstract</B></P> <P>The unique structure of kidney tubules is representative of their specialized function. Because maintaining tubular structure and controlled diameter is critical for kidney function, it is critical to understand how topographical cues, such as curvature, might alter cell morphology and biological characteristics. Here, we examined the effect of substrate curvature on the shape and phenotype of two kinds of renal epithelial cells (MDCK and HK-2) cultured on a microchannel with a broad range of principal curvature. We found that cellular architecture on curved substrates was closely related to the cell type-specific characteristics (stiffness, cell–cell adherence) of the cells and their density, as well as the sign and degree of curvature. As the curvature increased on convex channels, HK-2 cells, having lower cell stiffness and monolayer integrity than those of MDCK cells, aligned their in-plane axis perpendicular to the channel but did not significantly change in morphology. By contrast, MDCK cells showed minimal change in both morphology and alignment. However, on concave channels, both cell types were elongated and showed longitudinal directionality, although the changes in MDCK cells were more conservative. Moreover, substrate curvature contributed to cell polarization by enhancing the expression of apical and basolateral cell markers with height increase of the cells. Our study suggests curvature to be an important guiding principle for advanced tissue model developments, and that curved and geometrically ambiguous substrates can modulate the cellular morphology and phenotype.</P> <P><B>Statement of Significance</B></P> <P>In many tissues, such as renal tubules or intestinal villi, epithelial layers exist in naturally curved forms, a geometry that is not reproduced by flat cultures. Because maintaining tubular structure is critical for kidney function, it is important to understand how topographical cues, such as curvature, might alter cell morphology and biological characteristics. We found that cellular architecture on curved substrates was closely related to cell type and density, as well as the sign and degree of the curvature. Moreover, substrate curvature contributed to cell polarization by enhancing the expression of apical and basolateral cell markers with height increase. Our results suggested that substrate curvature might contribute to cellular architecture and enhance the polarization of kidney tubule cells.</P> <P><B>Graphical abstract</B></P> <P>[DISPLAY OMISSION]</P>

Induction of CD4+ Regulatory and Polarized Effector/helper T Cells by Dendritic Cells

Manfred B. Lutz 대한면역학회 2016 Immune Network Vol.16 No.1

Dendritic cells (DCs) are considered to play major roles during the induction of T cell immune responses as well as the maintenance of T cell tolerance. Naive CD4 + T cells have been shown to respond with high plasticity to signals inducing their polarization into effector/helper or regulatory T cells. Data obtained from in vitro generated bonemarrow (BM)-derived DCs as well as genetic mouse models revealed an important but not exclusive role of DCs in shaping CD4 + T cell responses. Besides the specialization of some conventional DC subsets for the induction of polarized immunity, also the maturation stage, activation of specialized transcription factors and the cytokine production of DCs have major impact on CD4 + T cells. Since in vitro generated BM-DCs show a high diversity to shape CD4 + T cells and their high similarity to monocyte-derived DCs in vivo, this review reports data mainly on BM-DCs in this process and only touches the roles of transcription factors or of DC subsets, which have been discussed elsewhere. Here, recent findings on 1) the conversion of naive into anergic and further into Foxp3 − regulatory T cells (Treg) by immature DCs, 2) the role of RelB in steady state migratory DCs (ssmDCs) for conversion of naive T cells into Foxp3 + Treg, 3) the DC maturation signature for polarized Th2 cell induction and 4) the DC source of IL-12 for Th1 induction are discussed.

The proteins encoded by the Drosophila Planar Polarity Effector genes inturned, fuzzy and fritz interact physically and can re-pattern the accumulation of ''upstream'' Planar Cell Polarity proteins

Wang, Y.,Yan, J.,Lee, H.,Lu, Q.,Adler, P.N. Academic Press 2014 Developmental Biology Vol.394 No.1

The frizzled/starry night pathway regulates planar cell polarity in a wide variety of tissues in many types of animals. It was discovered and has been most intensively studied in the Drosophila wing where it controls the formation of the array of distally pointing hairs that cover the wing. The pathway does this by restricting the activation of the cytoskeleton to the distal edge of wing cells. This results in hairs initiating at the distal edge and growing in the distal direction. All of the proteins encoded by genes in the pathway accumulate asymmetrically in wing cells. The pathway is a hierarchy with the Planar Cell Polarity (PCP) genes (aka the core genes) functioning as a group upstream of the Planar Polarity Effector (PPE) genes which in turn function as a group upstream of multiple wing hairs. Upstream proteins, such as Frizzled accumulate on either the distal and/or proximal edges of wing cells. Downstream PPE proteins accumulate on the proximal edge under the instruction of the upstream proteins. A variety of types of data support this hierarchy, however, we have found that when over expressed the PPE proteins can alter both the subcellular location and level of accumulation of the upstream proteins. Thus, the epistatic relationship is context dependent. We further show that the PPE proteins interact physically and can modulate the accumulation of each other in wing cells. We also find that over expression of Frtz results in a marked delay in hair initiation suggesting that it has a separate role/activity in regulating the cytoskeleton that is not shared by other members of the group.

Protein kinase C and calcineurin cooperatively mediate cell survival under compressive mechanical stress

Mishra, Ranjan,van Drogen, Frank,Dechant, Reinhard,Oh, Soojung,Jeon, Noo Li,Lee, Sung Sik,Peter, Matthias National Academy of Sciences 2017 PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF Vol.114 No.51

<P><B>Significance</B></P><P>Cells are constantly exposed to a variety of mechanical cues, and respond by activating conserved signaling pathways that modulate their profiliation and intracellular organization. For instance, cells in solid tumors experience sustained compression from the microenvironment, and compressive stress is known to trigger an invasive phenotype in some cancer cells. However, despite its importance for health and disease, the specific mechanosensors and the downstream signaling network mediating these physiological responses remain largely unknown, in part due to the lack of appropriate experimental systems. Here, we develop a microfluidic platform that allows triggering compressive mechanical stress in a reversible and controllable manner, and we identify cell surface receptors that specifically sense compressive mechanical stress and generate synergistic cellular responses.</P><P>Cells experience compressive stress while growing in limited space or migrating through narrow constrictions. To survive such stress, cells reprogram their intracellular organization to acquire appropriate mechanical properties. However, the mechanosensors and downstream signaling networks mediating these changes remain largely unknown. Here, we have established a microfluidic platform to specifically trigger compressive stress, and to quantitatively monitor single-cell responses of budding yeast in situ. We found that yeast senses compressive stress via the cell surface protein Mid2 and the calcium channel proteins Mid1 and Cch1, which then activate the Pkc1/Mpk1 MAP kinase pathway and calcium signaling, respectively. Genetic analysis revealed that these pathways work in parallel to mediate cell survival. Mid2 contains a short intracellular tail and a serine−threonine-rich extracellular domain with spring-like properties, and both domains are required for mechanosignaling. Mid2-dependent spatial activation of the Pkc1/Mpk1 pathway depolarizes the actin cytoskeleton in budding or shmooing cells, thereby antagonizing polarized growth to protect cells under compressive stress conditions. Together, these results identify a conserved signaling network responding to compressive mechanical stress, which, in higher eukaryotes, may ensure cell survival in confined environments.</P>

The Transcription Factor Mist1 Regulates the Cellular Polarity in Mouse Pancreatic Acinar Cells

Yu-Mi Yang,Syng-Ill Lee,Dong Min Shin KOREAN ACADAMY OF ORAL BIOLOGY 2012 International Journal of Oral Biology Vol.37 No.1

Pancreatic acinar cells exhibit a polarity that is characterized by the localization of secretory granules at the apical membrane. However, the factors that regulate cellular polarity in these cells are not well understood. In this study, we investigated the effect of Mist1, a basic helix-loop-helix transcription factor, on the cellular architecture of pancreatic acinar cells. Mist1-null mice displayed secretory granules that were diffuse throughout the pancreatic acinar cells, from the apical to basolateral membranes, whereas Mist1 heterozygote mice showed apical localization of secretory granules. Deletion of the Mist1 gene decreased the expression of type 3 inositol 1,4,5-triphosphate receptors (IP3R) but did not affect apical localization and expression of IP3R2. Mist1- null mice also displayed an increase in luminal areas and an increase in the expression of zymogen granules in pancreatic acinar cells. These results suggest that Mist1 plays a critical role in polar localization of cellular organelles and in maintaining cellular architecture in mouse pancreatic acinar cells.

PLEKHG3 enhances polarized cell migration by activating actin filaments at the cell front

Nguyen, Trang Thi Thu,Park, Wei Sun,Park, Byung Ouk,Kim, Cha Yeon,Oh, Yohan,Kim, Jin Man,Choi, Hana,Kyung, Taeyoon,Kim, Cheol-Hee,Lee, Gabsang,Hahn, Klaus M.,Meyer, Tobias,Heo, Won Do National Academy of Sciences 2016 PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF Vol.113 No.36

<P>Cells migrate by directing Ras-related C3 botulinum toxin substrate 1 (Rac1) and cell division control protein 42 (Cdc42) activities and by polymerizing actin toward the leading edge of the cell. Previous studies have proposed that this polarization process requires a local positive feedback in the leading edge involving Rac small GTPase and actin polymerization with PI3K likely playing a coordinating role. Here, we show that the pleckstrin homology and RhoGEF domain containing G3 (PLEKHG3) is a PI3K-regulated Rho guanine nucleotide exchange factor (RhoGEF) for Rac1 and Cdc42 that selectively binds to newly polymerized actin at the leading edge of migrating fibroblasts. Optogenetic inactivation of PLEKHG3 showed that PLEKHG3 is indispensable both for inducing and for maintaining cell polarity. By selectively binding to newly polymerized actin, PLEKHG3 promotes local Rac1/Cdc42 activation to induce more local actin polymerization, which in turn promotes the recruitment of more PLEKHG3 to induce and maintain cell front. Thus, autocatalytic reinforcement of PLEKHG3 localization to the leading edge of the cell provides a molecular basis for the proposed positive feedback loop that is required for cell polarization and directed migration.</P>

Optogenetic toolkit reveals the role of Ca²⁺ sparklets in coordinated cell migration

Kim, Jin Man,Lee, Minji,Kim, Nury,Heo, Won Do National Academy of Sciences 2016 PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF Vol.113 No.21

<P>Cell migration is controlled by various Ca2+ signals. Local Ca2+ signals, in particular, have been identified as versatile modulators of cell migration because of their spatiotemporal diversity. However, little is known about how local Ca2+ signals coordinate between the front and rear regions in directionally migrating cells. Here, we elucidate the spatial role of local Ca2+ signals in directed cell migration through combinatorial application of an optogenetic toolkit. An optically guided cell migration approach revealed the existence of Ca2+ sparklets mediated by L-type voltage-dependent Ca2+ channels in the rear part of migrating cells. Notably, we found that this locally concentrated Ca2+ influx acts as an essential transducer in establishing a global front-to-rear increasing Ca2+ gradient. This asymmetrical Ca2+ gradient is crucial for maintaining front-rear morphological polarity by restricting spontaneous lamellipodia formation in the rear part of migrating cells. Collectively, our findings demonstrate a clear link between local Ca2+ sparklets and front-rear coordination during directed cell migration.</P>

A novel Th1-type T-cell immunity-biasing effect of malate dehydrogenase derived from <i>Mycobacterium avium</i> subspecies <i>paratuberculosis</i> via the activation of dendritic cells

Kim, Woo Sik,Kim, Jong-Seok,Shin, Min-Kyoung,Shin, Sung Jae Elsevier 2018 Cytokine Vol.104 No.-

<P><B>Abstract</B></P> <P> <I>Mycobacterium avium</I> subspecies <I>paratuberculosis</I> (MAP) is the causative pathogen of Johne’s disease in ruminants, characterized by chronic granulomatous enteritis; it also has zoonotic potential and is associated with Crohn’s disease in humans. A better understanding of the mycobacterial antigens and their roles in the host immune response may facilitate the rational design of control strategies, including the development of effective vaccines and diagnostic tools. However, the functional roles of a large proportion of MAP antigens involved in modulating the host immune response remain unknown. In this study, an immunological role of MAP malate dehydrogenase (MDH, MAP2541c), an antigen that is upregulated in stress culture conditions, such as nutrient starvation and hypoxia, in polarizing naïve CD4<SUP>+</SUP>/CD8<SUP>+</SUP> T cells toward Th1-biased T-cell immunity via the activation of dendritic cells (DCs) was identified. DCs treated with MAP MDH displayed characteristics of the activated and mature immune status, with augmented expression of cell surface molecules and pro-inflammatory cytokines, including TNF-α, IL-1β, IL-6, and IL-12p70, but not IL-10, along with a dose-dependent decrease in the antigen uptake capacity. A mechanistic investigation revealed that the observed DC maturation is mediated by the activation of JNK, ERK, and p38 MAP kinases, and the NF-κB signaling pathway. Notably, DCs activated by MAP MDH treatment promoted naïve CD4<SUP>+</SUP>/CD8<SUP>+</SUP> T cell proliferation; in particular, they effectively polarized naïve CD4<SUP>+</SUP> T cells to secrete IFN-γ and IL-2 and activate T-bet, but, unlike the LPS control, did not influence IL-5 and GATA-3. These results indicated that MAP MDH has the potential to induce the Th1 cell response via DC activation. Collectively, our data demonstrated that MAP MDH is a novel immunostimulatory antigen that drives Th1-biased T cell polarization via interactions with DCs, suggesting that MDP MDH has the potential to be an effective MAP vaccine antigen target and diagnostic marker.</P> <P><B>Highlights</B></P> <P> <UL> <LI> MAP MDH activates DCs with increased expression of TNF-α, IL-1β, IL-6, and IL-12p70, but not IL-10. </LI> <LI> MAP MDH induces DC maturation by activation of NF-κB and MAPK. </LI> <LI> MAP MDH-treated DCs induce naïve CD4<SUP>+</SUP>/CD8<SUP>+</SUP> T cell proliferation and IFN-γ secretion in CD4<SUP>+</SUP> T cells. </LI> <LI> MAP MDH promotes Th1 type immunity via DC activation. </LI> </UL> </P>

TLR4-dependent effects of ISAg treatment on conventional T cell polarization in vivo

이성원,박현정,김서현,신수용,김경희,박상재,홍석만,전성호 한국통합생물학회 2019 Animal cells and systems Vol.23 No.3

We recently demonstrated that the polysaccharide component of the Korean medicinal herb Angelica gigas (immuno-stimulatory fraction of A. gigas; ISAg) induces anticancer effects in mice by activating natural killer (NK) and natural killer T (NKT) cells. However, it is unclear whether the use of ISAg in vivo can affect the differentiation of conventional T cells. Here, we investigated the effects of ISAg on the activation of conventional CD4+ and CD8+ T cells. We found that the administration of ISAg induced the polarization of CD4+ T cells toward the acquisition of the Th1 phenotype in vivo. Additionally, in mice treated with ISAg, CD8+ T cells produced more IFNγ than in control mice treated with PBS. Moreover, treatment with ISAg activated CD4+ and CD8+ T cells as well as NK and NKT cells, resulting in the secretion of Th1-type cytokines in a toll-like receptor 4 (TLR4)-dependent manner, implying that TLR4 is critical for an optimal Th1 response. Interestingly, ISAg treatment increased the number of Foxp3+ Treg cells, but not of Th2 cells, compared to control mice treated with PBS, indicating that ISAg possesses an immunomodulatory capacity that can control adaptive immune responses. Taken together, our results indicate that ISAg possesses a Th1-enhancing activity that could be used to treat Th2- mediated allergic immune diseases such as atopic dermatitis.