Cell Size Control Research Articles

PKA antagonizes CLASP-dependent microtubule stabilization to re-localize Pom1 and buffer cell size upon glucose limitation.

Cells couple growth with division and regulate size in response to nutrient availability. In rod-shaped fission yeast, cell-size control occurs at mitotic commitment. An important regulator is the DYRK-family kinase Pom1, which forms gradients from cell poles and inhibits the mitotic activator Cdr2, itself localized at the medial cortex. Where and when Pom1 modulates Cdr2 activity is unclear as Pom1 medial cortical levels remain constant during cell elongation. Here we show that Pom1 re-localizes to cell sides upon environmental glucose limitation, where it strongly delays mitosis. This re-localization is caused by severe microtubule destabilization upon glucose starvation, with microtubules undergoing catastrophe and depositing the Pom1 gradient nucleator Tea4 at cell sides. Microtubule destabilization requires PKA/Pka1 activity, which negatively regulates the microtubule rescue factor CLASP/Cls1/Peg1, reducing CLASP's ability to stabilize microtubules. Thus, PKA signalling tunes CLASP's activity to promote Pom1 cell side localization and buffer cell size upon glucose starvation.

Mathematical modeling reveals the mechanisms of feedforward regulation in cell fate decisions in budding yeast

The determination of cell fate is one of the key questions of developmental biology. Recent experiments showed that feedforward regulation is a novel feature of regulatory networks that controls reversible cellular transitions. However, the underlying mechanism of feedforward regulation‐mediated cell fate decision is still unclear. Therefore, using experimental data, we develop a full mathematical model of the molecular network responsible for cell fate selection in budding yeast. To validate our theoretical model, we first investigate the dynamical behaviors of key proteins at the Start transition point and the G1/S transition point; a crucial three‐node motif consisting of cyclin (Cln1/2), Substrate/Subunit Inhibitor of cyclin‐dependent protein kinase (Sic1) and cyclin B (Clb5/6) is considered at these points. The rapid switches of these important components between high and low levels at two transition check points are demonstrated reasonably by our model. Many experimental observations about cell fate decision and cell size control are also theoretically reproduced. Interestingly, the feedforward regulation provides a reliable separation between different cell fates. Next, our model reveals that the threshold for the amount of WHIskey (Whi5) removed from the nucleus is higher at the Reentry point in pheromone‐arrested cells compared with that at the Start point in cycling cells. Furthermore, we analyze the hysteresis in the cell cycle kinetics in response to changes in pheromone concentration, showing that Cln3 is the primary driver of reentry and Cln1/2 is the secondary driver of reentry. In particular, we demonstrate that the inhibition of Cln1/2 due to the accumulation of Factor ARrest (Far1) directly reinforces arrest. Finally, theoretical work verifies that the three‐node coherent feedforward motif created by cell FUSion (Fus3), Far1 and STErile (Ste12) ensures the rapid arrest and reversibility of a cellular state. The combination of our theoretical model and the previous experimental data contributes to the understanding of the molecular mechanisms of the cell fate decision at the G1 phase in budding yeast and will stimulate further biological experiments in future.

Evaluation of the Role of the opgGH Operon in Yersinia pseudotuberculosis and Its Deletion during the Emergence of Yersinia pestis.

The opgGH operon encodes glucosyltransferases that synthesize osmoregulated periplasmic glucans (OPGs) from UDP-glucose, using acyl carrier protein (ACP) as a cofactor. OPGs are required for motility, biofilm formation, and virulence in various bacteria. OpgH also sequesters FtsZ in order to regulate cell size according to nutrient availability. Yersinia pestis (the agent of flea-borne plague) lost the opgGH operon during its emergence from the enteropathogen Yersinia pseudotuberculosis. When expressed in OPG-negative strains of Escherichia coli and Dickeya dadantii, opgGH from Y. pseudotuberculosis restored OPGs synthesis, motility, and virulence. However, Y. pseudotuberculosis did not produce OPGs (i) under various growth conditions or (ii) when overexpressing its opgGH operon, its galUF operon (governing UDP-glucose), or the opgGH operon or Acp from E. coli. A ΔopgGH Y. pseudotuberculosis strain showed normal motility, biofilm formation, resistance to polymyxin and macrophages, and virulence but was smaller. Consistently, Y. pestis was smaller than Y. pseudotuberculosis when cultured at ≥ 37°C, except when the plague bacillus expressed opgGH. Y. pestis expressing opgGH grew normally in serum and within macrophages and was fully virulent in mice, suggesting that small cell size was not advantageous in the mammalian host. Lastly, Y. pestis expressing opgGH was able to infect Xenopsylla cheopis fleas normally. Our results suggest an evolutionary scenario whereby an ancestral Yersinia strain lost a factor required for OPG biosynthesis but kept opgGH (to regulate cell size). The opgGH operon was presumably then lost because OpgH-dependent cell size control became unnecessary.

EIF3 controls cell size independently of S6K1-activity.

All multicellular organisms require a life-long regulation of the number and the size of cells, which build up their organs. mTOR acts as a signaling nodule for the regulation of protein synthesis and growth. To activate the translational cascade, mTOR phosphorylates S6 kinase (S6K1), which is liberated from the eIF3-complex and mobilized for activation of its downstream targets. How S6K1 regulates cell size remains unclear. Here, we challenged cell size control through S6K1 by specifically depleting its binding partner eIF3 in normal and transformed cell lines. We show that loss of eIF3 leads to a massive reduction of cell size and cell number accompanied with an unexpected increase in S6K1-activity. The hyperactive S6K1-signaling was rapamycin-sensitive, suggesting an upstream mTOR-regulation. A selective S6K1 inhibitor (PF-4708671) was unable to interfere with the reduced size, despite efficiently inhibiting S6K1-activity. Restoration of eIF3 expression recovered size defects, without affecting the p-S6 levels. We further show that two, yet uncharacterized, cancer-associated mutations in the eIF3-complex, have the capacity to recover from reduced size phenotype, suggesting a possible role for eIF3 in regulating cancer cell size. Collectively, our results uncover a role for eIF3-complex in maintenance of normal and neoplastic cell size - independent of S6K1-signaling.

A noisy linear map underlies oscillations in cell size and gene expression in bacteria.

During bacterial growth, a cell approximately doubles in size before division, after which it splits into two daughter cells. This process is subjected to the inherent perturbations of cellular noise and thus requires regulation for cell-size homeostasis. The mechanisms underlying the control and dynamics of cell size remain poorly understood owing to the difficulty in sizing individual bacteria over long periods of time in a high-throughput manner. Here we measure and analyse long-term, single-cell growth and division across different Escherichia coli strains and growth conditions. We show that a subset of cells in a population exhibit transient oscillations in cell size with periods that stretch across several (more than ten) generations. Our analysis reveals that a simple law governing cell-size control-a noisy linear map-explains the origins of these cell-size oscillations across all strains. This noisy linear map implements a negative feedback on cell-size control: a cell with a larger initial size tends to divide earlier, whereas one with a smaller initial size tends to divide later. Combining simulations of cell growth and division with experimental data, we demonstrate that this noisy linear map generates transient oscillations, not just in cell size, but also in constitutive gene expression. Our work provides new insights into the dynamics of bacterial cell-size regulation with implications for the physiological processes involved.

Size sensors in bacteria, cell cycle control, and size control.

Bacteria proliferate by repetitive cycles of cellular growth and division. The progression into the cell cycle is admitted to be under the control of cell size. However, the molecular basis of this regulation is still unclear. Here I will discuss which mechanisms could allow coupling growth and division by sensing size and transmitting this information to the division machinery. Size sensors could act at different stages of the cell cycle. During septum formation, mechanisms controlling the formation of the Z ring, such as MinCD inhibition or Nucleoid Occlusion (NO) could participate in the size-dependence of the division process. In addition or alternatively, the coupling of growth and division may occur indirectly through the control of DNA replication initiation. The relative importance of these different size-sensing mechanisms could depend on the environmental and genetic context. The recent demonstration of an incremental strategy of size control in bacteria, suggests that DnaA-dependent control of replication initiation could be the major size control mechanism limiting cell size variation.

Phosphorylation of the exchange factor DENND3 by ULK in response to starvation activates Rab12 and induces autophagy.

Unc-51-like kinases (ULKs) are the most upstream kinases in the initiation of autophagy, yet the molecular mechanisms underlying their function are poorly understood. We report a new role for ULK in the induction of autophagy. ULK-mediated phosphorylation of the guanine nucleotide exchange factor DENND3 at serines 554 and 572 upregulates its GEF activity toward the small GTPase Rab12. Through binding to LC3 and associating with LC3-positive autophagosomes, active Rab12 facilitates autophagosome trafficking, thus establishing a crucial role for the ULK/DENND3/Rab12 axis in starvation-induced autophagy.

Slp2-a inactivates ezrin by recruiting protein phosphatase 1 to the plasma membrane

Synaptotagmin-like protein 2-a (Slp2-a) was originally described as a membrane trafficking protein that consists of a Slp homology domain (SHD), a linker domain, and tandem C2 domains (named the C2A domain and C2B domain). Slp2-a mediates docking of Rab27-bearing vesicles to the plasma membrane through simultaneous interaction with Rab27 and phospholipids in the plasma membrane. We have recently reported that Slp2-a regulates renal epithelial cell size through interaction with Rap1GAP2 via the C2B domain independently of Rab27 and demonstrated the presence of excess activation of ezrin, a membrane-cytoskeleton linker and signal transducer, in Slp2-a-knockdown Madin–Darby canine kidney II (MDCK II) cells. However, the precise mechanism of ezrin inactivation by Slp2-a in cell size control has remained largely unknown. In this study, we investigated the functional relationship between Slp2-a and ezrin in MDCK II cells. The results showed that activation of ezrin in control MDCK II cells either pharmacologically or by overexpression of a constitutively active ezrin mutant caused an increase in cell size, whereas inactivation of ezrin in Slp2-a-knockdown cells by a specific ezrin inhibitor restored them to their normal cell size. We also found that Slp2-a interacts via its previously uncharacterized linker domain with protein phosphatase 1β (PP1β), which inactivates ezrin, and that the interaction is required for the plasma membrane localization of PP1β. These results indicate that Slp2-a inactivates ezrin by recruiting PP1 to the plasma membrane.

The bacterial alarmone (p)ppGpp is required for virulence and controls cell size and survival of Pseudomonas syringae on plants.

The stringent response, mediated by second messenger (p)ppGpp, results in swift and massive transcriptional reprogramming under nutrient limited conditions. In this study, the role of (p)ppGpp on virulence of Pseudomonas syringae pv. syringae B728a (PssB728a) was investigated. The virulence of the relA/spoT (ppGpp(0) ) double mutant was completely impaired on bean, and bacterial growth was significantly reduced, suggesting that (p)ppGpp is required for full virulence of P. syringae. Expression of T3SS and other virulence genes was reduced in ppGpp(0) mutants. In addition, ppGpp deficiency resulted in loss of swarming motility, reduction of pyoverdine production, increased sensitivity to oxidative stress and antibiotic tolerance, as well as reduced ability to utilize γ-amino butyric acid. Increased levels of ppGpp resulted in reduced cell size of PssB728a when grown in a minimal medium and on plant surfaces, while most ppGpp(0) mutant cells were not viable on plant surfaces 24 h after spray inoculation, suggesting that ppGpp-mediated stringent response temporarily limits cell growth, and might control cell survival on plants by limiting their growth. These results demonstrated that ppGpp-mediated stringent response plays a central role in P. syringae virulence and survival and indicated that ppGpp serves as a global signal for regulating various virulence traits in PssB728a.

The necessary role of mTORC1 in central nervous system axon regeneration.

Permanent loss of vital functions after central nervous system (CNS) injury, e.g., blindness in traumatic optic nerve (ON) injury or paralysis in spinal cord injury, occurs in part because axons in the adult mammalian CNS do not regenerate after injury. Growth failure is due to the diminished intrinsic regenerative capacity of mature neurons and the inhibitory environment of the adult CNS. Neutralizing extracellular inhibitory molecules genetically or pharmacologically yields only limited regeneration and functional recovery, highlighting the critical importance of neuron-intrinsic factors. To explore the intrinsic regenerative signaling molecules, a relatively simple but robust in vivo model that replicates CNS traumatic injury and permits straightforward interpretation is required. Mouse retinal ganglion cell (RGC) and ON provide a valuable in vivo neural injury system to study intrinsic growth mechanisms in adult CNS neurons (Figure 1). Retina and ON are CNS structures that are essential for visual functions. RGC is the only neuronal type to relay visual information from retina to brain. The ON is formed by the projection axons sent exclusively from RGCs; it has the simplicity of a unidirectional axon pathway, which insures that any nerve fibers observed distal to a complete crush injury have regenerated and do not represent spared axons that underwent collateral sprouting or efferent axons from the brain to the retina. Its easy access allows adeno-associated viruses to be injected directly into the vitreous chamber of the eye and express transgenes specifically and efficiently in adult RGCs. This spatially and temporally controlled genetic manipulation allows us to overcome developmental issues associated with germ line manipulation and to test interventions that can potentially be translated to therapies. Exploiting the anatomical and technical advantages of the RGC/ON crush model to understand the intrinsic mechanisms of regenerative failure led us to the finding that deletion of phosphatase and tensin homolog (PTEN) promoted significant ON regeneration (Park et al., 2008). PTEN, a lipid phosphatase, is a major negative regulator of the phosphatidylinositol 3-kinase (PI3K)-mammalian target of rapamycin complex 1 (mTORC1) pathway. Similar axon regeneration phenotypes after PTEN deletion have been reported for mouse cortical motor neurons (Liu et al., 2010), drosophila sensory neurons (Song et al., 2012) and C. elegans motor neurons (Byrne et al., 2014), presumably through activating PI3K-mTORC1-controlled cell growth (Figure 2). Direct activation of mTORC1 also promotes axon regeneration in dopaminergic neurons (Kim et al., 2011) and peripheral nerves (Abe et al., 2010), further support for the critical role of mTORC1 in axon regeneration.

Mechanisms of daughter cell-size control during cell division.

Daughter cell size is tightly regulated during cell division. In animal cells, the position of the anaphase spindle specifies the cell cleavage site to dictate the relative size of the daughter cells. Although spindle orientation is regulated by dynein-dependent cortical pulling forces exerted on astral microtubules in many cell types, it was unclear how these forces are precisely regulated to center or displace the spindle. Recently, intrinsic signals derived from chromosomes or spindle poles have been demonstrated to regulate dynein-dependent pulling forces in symmetrically dividing cells. Unexpectedly, myosin-dependent contractile forces have also been shown to control spindle position by altering the cellular boundaries during anaphase. In this review, I discuss how dynein- and myosin-dependent forces are coordinately regulated to control daughter cell size.

Cell-Size Control and Homeostasis in Bacteria

How cells control their size and maintain size homeostasis is a fundamental open question. Cell-size homeostasis has been discussed in the context of two major paradigms: "sizer," in which the cell actively monitors its size and triggers the cell cycle once it reaches a critical size, and "timer," in which the cell attempts to grow for a specific amount of time before division. These paradigms, in conjunction with the "growth law" [1] and the quantitative bacterial cell-cycle model [2], inspired numerous theoretical models [3-9] and experimental investigations, from growth [10, 11] to cell cycle and size control [12-15]. However, experimental evidence involved difficult-to-verify assumptions or population-averaged data, which allowed different interpretations [1-5, 16-20] or limited conclusions [4-9]. In particular, population-averaged data and correlations are inconclusive as the averaging process masks causal effects at the cellular level. In this work, we extended a microfluidic "mother machine" [21] and monitored hundreds of thousands of Gram-negative Escherichia coli and Gram-positive Bacillus subtilis cells under a wide range of steady-state growth conditions. Our combined experimental results and quantitative analysis demonstrate that cells add a constant volume each generation, irrespective of their newborn sizes, conclusively supporting the so-called constant Δ model. This model was introduced for E. coli [6, 7] and recently revisited [9], but experimental evidence was limited to correlations. This "adder" principle quantitatively explains experimental data at both the population and single-cell levels, including the origin and the hierarchy of variability in the size-control mechanisms and how cells maintain size homeostasis.

A Constant Size Extension Drives Bacterial Cell Size Homeostasis

Cell size control is an intrinsic feature of the cell cycle. In bacteria, cell growth and division are thought to be coupled through a cell size threshold. Here, we provide direct experimental evidence disproving the critical size paradigm. Instead, we show through single-cell microscopy and modeling that the evolutionarily distant bacteria Escherichia coli and Caulobacter crescentus achieve cell size homeostasis by growing, on average, the same amount between divisions, irrespective of cell length at birth. This simple mechanism provides a remarkably robust cell size control without the need of being precise, abating size deviations exponentially within a few generations. This size homeostasis mechanism is broadly applicable for symmetric and asymmetric divisions, as well as for different growth rates. Furthermore, our data suggest that constant size extension is implemented at or close to division. Altogether, our findings provide fundamentally distinct governing principles for cell size and cell-cycle control in bacteria.

Mutations in PTRH2 cause novel infantile‐onset multisystem disease with intellectual disability, microcephaly, progressive ataxia, and muscle weakness

ObjectiveTo identify the cause of a so-far unreported phenotype of infantile-onset multisystem neurologic, endocrine, and pancreatic disease (IMNEPD).MethodsWe characterized a consanguineous family of Yazidian-Turkish descent with IMNEPD. The two affected children suffer from intellectual disability, postnatal microcephaly, growth retardation, progressive ataxia, distal muscle weakness, peripheral demyelinating sensorimotor neuropathy, sensorineural deafness, exocrine pancreas insufficiency, hypothyroidism, and show signs of liver fibrosis. We performed whole-exome sequencing followed by bioinformatic analysis and Sanger sequencing on affected and unaffected family members. The effect of mutations in the candidate gene was studied in wild-type and mutant mice and in patient and control fibroblasts.ResultsIn a consanguineous family with two individuals with IMNEPD, we identified a homozygous frameshift mutation in the previously not disease-associated peptidyl-tRNA hydrolase 2 (PTRH2) gene. PTRH2 encodes a primarily mitochondrial protein involved in integrin-mediated cell survival and apoptosis signaling. We show that PTRH2 is highly expressed in the developing brain and is a key determinant in maintaining cell survival during human tissue development. Moreover, we link PTRH2 to the mTOR pathway and thus the control of cell size. The pathology suggested by the human phenotype and neuroimaging studies is supported by analysis of mutant mice and patient fibroblasts.InterpretationWe report a novel disease phenotype, show that the genetic cause is a homozygous mutation in the PTRH2 gene, and demonstrate functional effects in mouse and human tissues. Mutations in PTRH2 should be considered in patients with undiagnosed multisystem neurologic, endocrine, and pancreatic disease.

Subcellular localization of era and their effects on cell size

s for the 29th Annual Congress of Japanese Society / Journal of Reproductive Immunology 106 (2014) 1–20 9 each patient. Among RPL patients, 90 patients (30%) were deficient for total protein S antigen (<65%). Only 8 patients out of these 90 patients were also deficient for protein S activity (<60%). Fifty-seven (27.1%) out of 210 anti-protein S antibody negative patients, 20 (36.4%) out of 55 antiprotein S antibody positive (1+ ormore) patients, 6 (54.5%) out of 11 positive (2+ or more) patients, 4 (100%) out of 4 positive (3+) patients were protein S deficient. There was a significant correlation between total protein S antigen level and autoantibodies to protein S (p=0.0061, OR 24.03). Among 20 protein S deficient patients who were positive (1+ or more) for anti-protein S antibodies and 61 protein S deficient patients who were negative for anti-protein S antibodies, the incidence of each factor was as follows; aPTTmixing test20%vs3.3% (p=0.0304,OR7.38), antiphosphatidylethanolamine antibodies 35% vs 9.8% (p=0.0137, OR 4.94), FXII deficiency 50% vs 23% (p=0.0274, OR 3.36), anti-prothrombin/phoshpatidylserine antibodies 30% vs 18% (NS), anti-FXII antibodies 30% vs 21.3% (NS), anticardiolipin antibodies 5.0% vs 6.6% (NS), respectively. These data indicate a correlation between anti-protein S antibodies and autoantibodies to contact proteins (e.g. FXII and kininogens). Protein Smay play an important role in reproduction. http://dx.doi.org/10.1016/j.jri.2014.09.020 A case report of neonatal hemochromatosis treated by high-dose intravenous immunoglobulin K. Motomura1,4,∗, A. Sasaki1, M. Hisano1, K. Yamaguchi1, Y. Ito1, R. Ito2, M. Kasahara3, K. Matsumoto4, H. Sago1 1 Center of Maternal-Fetal, Neonatal and Reproductive Medicine, National Center for Child Health and Development, Japan 2 Department of Medical Subspecialties, National Center for Child Health and Development, Japan 3 Organ Transplantation Center, National Center for Child Health and Development, Japan 4 Department of Allergy and Immunology, National Research Institute for Child Heath and Development, Japan Neonatal hemochromatosis (NH) is characterized by serious fetal or neonatal liver failure due to alloimmune liver damage. Specific diagnosis using immunohistochemical staining ofmembrane attack complex (MAC IHC) in the liver and maternal treatment with high-dose intravenous immunoglobulin (IVIG) have recently become the standard management. Here, we report a successful pregnancy in a woman whose index pregnancy was diagnosed as NH and who was treated with IVIG. A 37-year-old, gravida 1 para 0 woman delivered a normal-appearing neonate who died of liver failure only 17h after delivery in spite of having shown no symptoms during gestation or delivery. The neonate was diagnosed as NH based on positive MAC IHC staining. After this pregnancy, themother experienced two spontaneousmiscarriages.Duringher latestpregnancy, she received IVIG (1 g/kg/week) after 14 weeks of gestation. She had no complications, and delivered a healthy neonate at term. The neonate showed slight abnormalities in liver function and iron metabolism, which normalized without any treatment. In conclusion, MAC IHCwas useful for diagnosis, and IVIG markedly improved the outcome of the neonate. http://dx.doi.org/10.1016/j.jri.2014.09.021 Subcellular localization of era and their effects on cell size Z.-L. Li ∗, Y. Otsuki Department of Anatomy & Biology, Osaka Medical College, Japan Current data support the idea that estrogen receptor alpha (ER ) is located in cell membrane, cytoplasm, and/or nucleus. ER plays important roles in such processes as cellular proliferation, differentiation, migration and invasion respectively or collectively. In these processes, ER cross-talkwithother factors to control distinctive groupsof target genes genetically and epigenetically. While some of these factors appear to regulate the accessibility of promoters, others are members of various kinase cascades such as PI3K-AKT that control crucial cellular events. Although some of these factors have been implicated in regulation of cell size, the function of ER in this aspect is unclear. In order to discern functions and interactions of ER and its co-regulators, we have allocated ER to different locations in ER -negative Ishikawa cell, using permanent transfection technique. In this report, wewould present the results showing how ER in different subcellular locations control cell size. Our study might shed light on the signalling pathway down-stream of ER that regulate cell size. http://dx.doi.org/10.1016/j.jri.2014.09.022 A new local drug delivery system for the uterus using bio-nanocapsule (BNC) K. Koizumi1,∗, H. Nakamura1, T. Matsuzaki2, S. Kuroda3, T. Takiuchi1, K. Kumasawa1, T. Kimura1 1 Department of Obstetrics and Gynaecololgy, Osaka University Graduate School of Medicine, Japan 2 Department of Cardiovascular Medicine, Osaka University Graduate School of Medicine, Japan 3 Laboratory of industrial Biosciences, Division of Biotechnology, Department of Bioengineering Sciences, Graduate School of Bioagricultural Sciences, Nagoya University,

A case report of neonatal hemochromatosis treated by high-dose intravenous immunoglobulin

s for the 29th Annual Congress of Japanese Society / Journal of Reproductive Immunology 106 (2014) 1–20 9 each patient. Among RPL patients, 90 patients (30%) were deficient for total protein S antigen (<65%). Only 8 patients out of these 90 patients were also deficient for protein S activity (<60%). Fifty-seven (27.1%) out of 210 anti-protein S antibody negative patients, 20 (36.4%) out of 55 antiprotein S antibody positive (1+ ormore) patients, 6 (54.5%) out of 11 positive (2+ or more) patients, 4 (100%) out of 4 positive (3+) patients were protein S deficient. There was a significant correlation between total protein S antigen level and autoantibodies to protein S (p=0.0061, OR 24.03). Among 20 protein S deficient patients who were positive (1+ or more) for anti-protein S antibodies and 61 protein S deficient patients who were negative for anti-protein S antibodies, the incidence of each factor was as follows; aPTTmixing test20%vs3.3% (p=0.0304,OR7.38), antiphosphatidylethanolamine antibodies 35% vs 9.8% (p=0.0137, OR 4.94), FXII deficiency 50% vs 23% (p=0.0274, OR 3.36), anti-prothrombin/phoshpatidylserine antibodies 30% vs 18% (NS), anti-FXII antibodies 30% vs 21.3% (NS), anticardiolipin antibodies 5.0% vs 6.6% (NS), respectively. These data indicate a correlation between anti-protein S antibodies and autoantibodies to contact proteins (e.g. FXII and kininogens). Protein Smay play an important role in reproduction. http://dx.doi.org/10.1016/j.jri.2014.09.020 A case report of neonatal hemochromatosis treated by high-dose intravenous immunoglobulin K. Motomura1,4,∗, A. Sasaki1, M. Hisano1, K. Yamaguchi1, Y. Ito1, R. Ito2, M. Kasahara3, K. Matsumoto4, H. Sago1 1 Center of Maternal-Fetal, Neonatal and Reproductive Medicine, National Center for Child Health and Development, Japan 2 Department of Medical Subspecialties, National Center for Child Health and Development, Japan 3 Organ Transplantation Center, National Center for Child Health and Development, Japan 4 Department of Allergy and Immunology, National Research Institute for Child Heath and Development, Japan Neonatal hemochromatosis (NH) is characterized by serious fetal or neonatal liver failure due to alloimmune liver damage. Specific diagnosis using immunohistochemical staining ofmembrane attack complex (MAC IHC) in the liver and maternal treatment with high-dose intravenous immunoglobulin (IVIG) have recently become the standard management. Here, we report a successful pregnancy in a woman whose index pregnancy was diagnosed as NH and who was treated with IVIG. A 37-year-old, gravida 1 para 0 woman delivered a normal-appearing neonate who died of liver failure only 17h after delivery in spite of having shown no symptoms during gestation or delivery. The neonate was diagnosed as NH based on positive MAC IHC staining. After this pregnancy, themother experienced two spontaneousmiscarriages.Duringher latestpregnancy, she received IVIG (1 g/kg/week) after 14 weeks of gestation. She had no complications, and delivered a healthy neonate at term. The neonate showed slight abnormalities in liver function and iron metabolism, which normalized without any treatment. In conclusion, MAC IHCwas useful for diagnosis, and IVIG markedly improved the outcome of the neonate. http://dx.doi.org/10.1016/j.jri.2014.09.021 Subcellular localization of era and their effects on cell size Z.-L. Li ∗, Y. Otsuki Department of Anatomy & Biology, Osaka Medical College, Japan Current data support the idea that estrogen receptor alpha (ER ) is located in cell membrane, cytoplasm, and/or nucleus. ER plays important roles in such processes as cellular proliferation, differentiation, migration and invasion respectively or collectively. In these processes, ER cross-talkwithother factors to control distinctive groupsof target genes genetically and epigenetically. While some of these factors appear to regulate the accessibility of promoters, others are members of various kinase cascades such as PI3K-AKT that control crucial cellular events. Although some of these factors have been implicated in regulation of cell size, the function of ER in this aspect is unclear. In order to discern functions and interactions of ER and its co-regulators, we have allocated ER to different locations in ER -negative Ishikawa cell, using permanent transfection technique. In this report, wewould present the results showing how ER in different subcellular locations control cell size. Our study might shed light on the signalling pathway down-stream of ER that regulate cell size. http://dx.doi.org/10.1016/j.jri.2014.09.022 A new local drug delivery system for the uterus using bio-nanocapsule (BNC) K. Koizumi1,∗, H. Nakamura1, T. Matsuzaki2, S. Kuroda3, T. Takiuchi1, K. Kumasawa1, T. Kimura1 1 Department of Obstetrics and Gynaecololgy, Osaka University Graduate School of Medicine, Japan 2 Department of Cardiovascular Medicine, Osaka University Graduate School of Medicine, Japan 3 Laboratory of industrial Biosciences, Division of Biotechnology, Department of Bioengineering Sciences, Graduate School of Bioagricultural Sciences, Nagoya University,

Principles of Bacterial Cell-Size Determination Revealed by Cell-Wall Synthesis Perturbations

Although bacterial cell morphology is tightly controlled, the principles of size regulation remain elusive. In Escherichia coli, perturbation of cell-wall synthesis often results in similar morphologies, making it difficult to deconvolve the complex genotype-phenotype relationships underlying morphogenesis. Here we modulated cell width through heterologous expression of sequences encoding the essential enzyme PBP2 and through sublethal treatments with drugs that inhibit PBP2 and the MreB cytoskeleton. We quantified the biochemical and biophysical properties of the cell wall across a wide range of cell sizes. We find that, although cell-wall chemical composition is unaltered, MreB dynamics, cell twisting, and cellular mechanics exhibit systematic large-scale changes consistent with altered chirality and a more isotropic cell wall. This multiscale analysis enabled identification of distinct roles for MreB and PBP2, despite having similar morphological effects when depleted. Altogether, our results highlight the robustness of cell-wall synthesis and physical principles dictating cell-size control.

A Genomic Multiprocess Survey of Machineries that Control and Link Cell Shape, Microtubule Organization, and Cell-Cycle Progression

Understanding cells as integrated systems requiresthat we systematically decipher how single genesaffect multiple biological processes and how processes are functionally linked. Here, we used multiprocess phenotypic profiling, combining high-resolution 3D confocal microscopy and multiparametric image analysis, to simultaneously survey the fission yeast genome with respect to three key cellular processes: cell shape, microtubule organization, and cell-cycle progression. We identify, validate, and functionally annotate 262 genes controlling specific aspects of those processes. Of these, 62% had not been linked to these processes before and 35% are implicated in multiple processes. Importantly, we identify a conserved role for DNA-damage responses in controlling microtubule stability. In addition, we investigate how the processes are functionally linked. We show unexpectedly that disruption of cell-cycle progression does not necessarily affect cell size control and that distinct aspects of cell shape regulate microtubules and vice versa, identifying important systems-level links across these processes.

Abstract 2253: YAP nuclear accumulation involved in drug-induced cellular senescence in vitro

Abstract Cellular senescence is accompanied by some striking morphological changes, such as becoming large, flat and multinucleated. However, how these changes are established is poorly understood. The Hippo pathway plays an important role in the control of organ size. Previous study reported that YAP, one major downstream effector of Hippo pathway, could be regulated by mechanical signals through F-actin bundles. However, whether YAP participates in the control of cell size through cytoskeleton remains unexplained. In our current study, we found that the senescent cells induced with adriamycin or camptothecin became large and flat, and the stress fibers of the senescent cells seemed to maintain or increase, observed by F-actin staining, though in a way of inappropriate actin clumping. Moreover increased nuclear level and activity of YAP were identified in drug-induced senescent cells. Hence, our data indicated that the increased F-actin bundles and mechanical signals could be acquired in senescent cells induced by adriamycin or camptothecin. In summary, based on our findings here, we proposed that YAP might play some roles during drug-induced cellular senescence in vitro. Citation Format: Kai Ma, Shuren Wang, Qing Xu, Mei Liu, Hongxia Zhu, Ningzhi Xu. YAP nuclear accumulation involved in drug-induced cellular senescence in vitro. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2253. doi:10.1158/1538-7445.AM2014-2253

Metabolic pathways further increase the complexity of cell size control in budding yeast.

How organisms regulate their size is a major question in biology. With a few notable exceptions (such as cell divisions in the early embryo), most cells need to reach a critical size in order to initiate a new cell cycle. How cells set a critical cell size, and how they know it has been reached, is not well understood. Using various types of experimental systems, decades ago two main models were proposed for cell size homeostasis: the deterministic model and the probabilistic model.