• Molecules and Cells
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• 제39권4호
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• pp.322-329
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• 2016

Voltage-gated $Ca^{2+}$ ($Ca_V$) channels are dynamically modulated by Gprotein-coupled receptors (GPCR). The $M_1$ muscarinic receptor stimulation is known to enhance $Ca_V2.3$ channel gating through the activation of protein kinase C (PKC). Here, we found that $M_1$ receptors also inhibit $Ca_V2.3$ currents when the channels are fully activated by PKC. In whole-cell configuration, the application of phorbol 12-myristate 13-acetate (PMA), a PKC activator, potentiated $Ca_V2.3$ currents by ~two-fold. After the PMA-induced potentiation, stimulation of $M_1$ receptors decreased the $Ca_V2.3$ currents by $52{\pm}8%$. We examined whether the depletion of phosphatidylinositol 4,5-bisphosphate ($PI(4,5)P_2$) is responsible for the muscarinic suppression of $Ca_V2.3$ currents by using two methods: the Danio rerio voltage-sensing phosphatase (Dr-VSP) system and the rapamycin-induced translocatable pseudojanin (PJ) system. First, dephosphorylation of $PI(4,5)P_2$ to phosphatidylinositol 4-phosphate (PI(4)P) by Dr-VSP significantly suppressed $Ca_V2.3$ currents, by $53{\pm}3%$. Next, dephosphorylation of both PI(4)P and $PI(4,5)P_2$ to PI by PJ translocation further decreased the current by up to $66{\pm}3%$. The results suggest that $Ca_V2.3$ currents are modulated by the $M_1$ receptor in a dual mode-that is, potentiation through the activation of PKC and suppression by the depletion of membrane $PI(4,5)P_2$. Our results also suggest that there is rapid turnover between PI(4)P and $PI(4,5)P_2$ in the plasma membrane.

• Molecules and Cells
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• 제22권1호
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• pp.97-103
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• 2006

Phosphoinositides are critical regulators of ion channel and transporter activity. There are multiple isomers of biologically active phosphoinositides in the plasma membrane and the different lipid species are non-randomly distributed. However, the mechanism by which cells impose selectivity and directionality on lipid movements and so generate a non-random lipid distribution remains unclear. In the present study we investigated which structural elements of phosphoinositides are responsible for their subcellular location and movement. We incubated phosphatidylinositol (PI), phosphatidylinositol 4-monophosphate (PI(4)P) and phosphatidylinositol 4,5-bisphosphate ($PI(4,5)P_2$) with short or long acyl chains in CHO and HEK cells. We show that phosphate number and acyl chain length determine cellular location and translocation movement. In CHO cells, $PI(4,5)P_2$ with a long acyl chain was released into the cytosol easily because of a low partition coefficient whereas long chain PI was released more slowly because of a high partition coefficient. In HEK cells, the cellular location and translocation movement of PI were similar to those of PI in CHO cells, whereas those of $PI(4,5)P_2$ were different; some mechanism restricted the translocation movement of $PI(4,5)P_2$, and this is in good agreement with the extremely low lateral diffusion of $PI(4,5)P_2$. In contrast to the dependence on the number of phosphates of the phospholipid head group of long acyl chain analogs, short acyl chain phospholipids easily undergo translocation movement regardless of cell type and number of phosphates in the lipid headgroup.

• 한국약용작물학회:학술대회논문집
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• 한국약용작물학회 2011년도 추계학술발표회
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• pp.31-45
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• 2011

The amyloid ${\beta}$-peptide ($A{\beta}$), which originates from the proteolytic cleavage of amyloid precursor protein (APP), plays a central role in the pathogenesis of Alzheimer's disease (AD). Mounting evidence indicates that different species of $A{\beta}$, such as $A{\beta}$ oligomers and fibrils, may contribute to AD pathogenesis via distinct mechanisms at different stages of the disease. Importantly, elevation and accumulation of soluble $A{\beta}$ oligomers closely correlate with cognitive decline and/or disease progression in animal models of AD. In agreement with these studies, oligomers of $A{\beta}$ have been shown to directly affect synaptic plasticity, a neuronal process that is known to be essential for memory formation. Our previous studies showed that $A{\beta}$ induces the breakdown of phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2), a phospholipid that regulates key aspects of neuronal function. PI(4,5)P2 breakdown was found to be a key step toward synaptic and memory dysfunction in a mouse model of AD. To this end, we seek to identify small molecules that could elevate the levels of PI(4,5)P2 and subsequently block $A{\beta}$ oligomer-induced breakdown of PI(4,5)P2 and synaptic dysfunction.. We found that (20S)Rg3, an active triterpene glycoside from heat-processed ginseng, serves as an agonist for phosphatidylinositol 4-kinase IIalpha (PI4KIIalpha), which is a lipid kinase that mediates a rate-limiting step in PI(4,5)P2 synthesis. Consequently, (20S)Rg3 stimulates PI(4,5)P2 synthesis by directly stimulating the activity of PI4KIIalpha. Interestingly, treatment of a mouse model of AD with (20S)Rg3 leads to reversal of memory deficits. Our data suggest that the PI(4,5)P2-promoting effects of (20S)Rg3 may help mitigate the cognitive symptoms associated with AD.

• Korean Journal of Radiology
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• 제22권7호
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• pp.1100-1109
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• 2021

Objective: To compare the diagnostic performance between Prostate Imaging-Reporting and Data System version 2.0 (PI-RADSv2.0) and version 2.1 (PI-RADSv2.1) for clinically significant prostate cancer (csPCa) in the peripheral zone (PZ). Materials and Methods: This retrospective study included 317 patients who underwent multiparametric magnetic resonance imaging and targeted biopsy for PZ lesions. Definition of csPCa was International Society of Urologic Pathology grade ≥ 2 cancer. Area under the curve (AUC), sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), and accuracy for csPCa were analyzed by two readers. The cancer detection rate (CDR) for csPCa was investigated according to the PI-RADS categories. Results: AUC of PI-RADSv2.1 (0.856 and 0.858 for reader 1 and 2 respectively) was higher than that of PI-RADSv2.0 (0.795 and 0.747 for reader 1 and 2 respectively) (both p < 0.001). Sensitivity, specificity, PPV, NPV, and accuracy for PI-RADSv2.0 vs. PI-RADSv2.1 were 93.2% vs. 88.3% (p = 0.023), 52.8% vs. 76.6% (p < 0.001), 48.7% vs. 64.5% (p < 0.001), 94.2% vs. 93.2% (p = 0.504), and 65.9% vs. 80.4% (p < 0.001) for reader 1, and 96.1% vs. 92.2% (p = 0.046), 34.1% vs. 72.4% (p < 0.001), 41.3% vs. 61.7% (p < 0.001), 94.8% vs. 95.1% (p = 0.869), and 54.3% vs. 78.9% (p < 0.001) for reader 2, respectively. CDRs of PI-RADS categories 1-2, 3, 4, and 5 for PI-RADSv2.0 vs. PI-RADSv2.1 were 5.9% vs. 5.9%, 5.8% vs. 12.5%, 39.8% vs. 56.2%, and 88.9% vs. 88.9% for reader 1; and 4.5% vs. 4.1%, 6.1% vs. 11.1%, 32.5% vs. 53.4%, and 85.0% vs. 86.8% for reader 2, respectively. Conclusion: Our data demonstrated improved AUC, specificity, PPV, accuracy, and CDRs of category 3 or 4 of PI-RADSv2.1, but decreased sensitivity, compared with PI-RADSv2.0, for csPCa in PZ.

• BMB Reports
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• 제47권7호
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• pp.361-368
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• 2014

Lipid components in biological membranes are essential for maintaining cellular function. Phosphoinositides, the phosphorylated derivatives of phosphatidylinositol (PI), regulate many critical cell processes involving membrane signaling, trafficking, and reorganization. Multiple metabolic pathways including phosphoinositide kinases and phosphatases and phospholipases tightly control spatio-temporal concentration of membrane phosphoinositides. Metabolizing enzymes responsible for PI 4,5-bisphosphate (PI(4,5)P2) production or degradation play a regulatory role in Toll-like receptor (TLR) signaling and trafficking. These enzymes include PI 4-phosphate 5-kinase, phosphatase and tensin homolog, PI 3-kinase, and phospholipase C. PI(4,5)P2 mediates the interaction with target cytosolic proteins to induce their membrane translocation, regulate vesicular trafficking, and serve as a precursor for other signaling lipids. TLR activation is important for the innate immune response and is implicated in diverse pathophysiological disorders. TLR signaling is controlled by specific interactions with distinct signaling and sorting adaptors. Importantly, TLR signaling machinery is differentially formed depending on a specific membrane compartment during signaling cascades. Although detailed mechanisms remain to be fully clarified, phosphoinositide metabolism is promising for a better understanding of such spatio-temporal regulation of TLR signaling and trafficking.

• BMB Reports
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• 제54권6호
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• pp.311-316
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• 2021

Ethanol often causes critical health problems by altering the neuronal activities of the central and peripheral nerve systems. One of the cellular targets of ethanol is the plasma membrane proteins including ion channels and receptors. Recently, we reported that ethanol elevates membrane excitability in sympathetic neurons by inhibiting Kv7.2/7.3 channels in a cell type-specific manner. Even though our studies revealed that the inhibitory effects of ethanol on the Kv7.2/7.3 channel was diminished by the increase of plasma membrane phosphatidylinositol 4,5-bisphosphate (PI(4,5)P₂), the molecular mechanism of ethanol on Kv7.2/7.3 channel inhibition remains unclear. By investigating the kinetics of Kv7.2/7.3 current in high K⁺ solution, we found that ethanol inhibited Kv7.2/7.3 channels through a mechanism distinct from that of tetraethylammonium (TEA) which enters into the pore and blocks the gate of the channels. Using a non-stationary noise analysis (NSNA), we demonstrated that the inhibitory effect of ethanol is the result of reduction of open probability (P_O) of the Kv7.2/7.3 channel, but not of a single channel current (i) or channel number (N). Finally, ethanol selectively facilitated the kinetics of Kv7.2 current suppression by voltage-sensing phosphatase (VSP)-induced PI(4,5)P₂ depletion, while it slowed down Kv7.2 current recovery from the VSP-induced inhibition. Together our results suggest that ethanol regulates neuronal activity through the reduction of open probability and PI(4,5)P₂ sensitivity of Kv7.2/7.3 channels.

• 수면정신생리
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• 제19권2호
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• pp.68-76
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• 2012

목 적: 수면 장애는 자율신경계의 변화를 유발하여 심혈관계에 영향을 준다. 일차성 불면증(primary insomnia, 이하 PI)은 수면부족 및 각성으로, 폐쇄성 수면무호흡 증후군(obstructive sleep apnea syndrome, 이하 OSAS)은 수면 중 빈번한 각성, 저산소증 등으로 교감신경계 항진을 유발한다. 이에 OSAS와 PI 사이에서 심박동률 변이도(heart rate variability, 이하 HRV) 분석을 통해 자율신경계의 변화 정도를 비교하고자 하였다. 방 법: 임상병력 및 야간수면다원검사(nocturnal polysomnography, 이하 NPSG)로 선택된 PI와 OSAS 전체 315 명의 대상자 중 OSAS는 무호흡-저호흡 지수(apnea-hypopnea index, 이하 AHI)에 따라서 경도, 중등도, 중증도 OSAS로 분류하여 PI를 포함하여 4개의 군으로 분류되었다. 이후 연령, 성별, 군간 개체수를 고려하여 최종적으로 110명의 대상자가 선택되었고, PI군(평균나이 41.50, 표준편차 13.16, AHI <5, n=20), 경도 OSAS군(평균나이 43.67, 표준편차 12.11, AHI 5~15, n=30), 중등도 OSAS군(평균나이 44.93, 표준편차 12.38, AHI 16~30, n=30), 중증도 OSAS군(평균 나이 45.87, 표준편차 12.44, AHI >30, n=30)의 4개 군으로 나누었다. 이후 NPSG를 통해 얻은 각 군의 HRV 지수 비교를 위하여 연령과 체질량지수(body weight index, BMI)를 공변량으로 하여 공분산분석(analysis of covariance, ANCOVA)를 실시한 후 사후분석으로 Sidak test를 실시하였다. 결 과: HRV 지수의 비교 결과 PI군과 경도, 중등도의 OSAS군 간에 유의한 차이는 없었다. 그러나 PI군과 중증도 OSAS군 사이에는 유의한 차이가 있었다. 군간비교에서 통계적으로 가장 현저한 차이를 보인 PI군과 중증도 OSAS군에서 HRV 지수 중 통계적으로 의미 있는 각각의 값은 다음과 같았다. Average RR interval은 $991.1{\pm}27.1$과 $875.8{\pm}22.0$ ms(p=0.016), standard deviation of NN interval(SDNN)은 $85.4{\pm}6.6$과 $112.8{\pm}5.4$ ms(p=0.022), SDNN index는 $57.5{\pm}5.2$과 $87.6{\pm}4.2$(p<0.001), total power는 $11893.5{\pm}1359.9$과 $8097.0{\pm}1107.2ms^2$(p=0.008), very low frequency(VLF)는 $7534.8{\pm}1120.1$과 $11883.8{\pm}912.0ms^2$(p=0.035), low frequency(LF)는 $2724.2{\pm}327.8$과 $4351.6{\pm}266.9ms^2$였다(p=0.003). 결 론: 본 연구에서는 PI군에 비해 중증도의 OSAS군에서 HRV 지수 중 교감신경계의 영향을 많이 받는 VLF, LF 등이 상승하는 양상을 보였고, 이를 통해 PI에 비해 중증도의 OSAS가 교감신경계 활동증가에 보다 더 큰 영향을 끼치는 것을 알 수 있었다.

• 식물조직배양학회지
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• 제25권1호
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• pp.45-50
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• 1998

꽃양배추의 하배축 조직을 proteinase inhibitor II 유전자가 도입된 Agrobacterium tumefaciens LBA 4404와 2일간 pH 5.5로 조절된 MS 액체배지에서 공동배양후 carbenicillin 500mg/L kanamycin 20mg/L와 BA 1mg/L가 함유된 MS 재분화배지에 옮겼다. 이들 조직을 매 2주마다 계대배양하였으며 약 4주후에 kanamycin 저항성 개체를 얻었다. 형질전환된 것으로 추정되는 식물체는 kanamycin 30mg/L가 함유된 선발배지에서 생존하였다. PCR 분석결과, PI-II 유전자가 형질전환체의 게놈상에 삽입되어 있음을 확인하였다. 형질전환체의 Southern blot 분석을 통하여 ECL-labelling된 PI-II 유전자와 동일한 것으로 판단되는 약 500bp 위치에서 밴드를 확인할 수 있었다.

• 생명과학회지
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• 제26권8호
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• pp.875-880
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• 2016

진핵세포의 원형질막은 외부환경으로부터 세포를 격리하는 물리적인 장벽뿐만 아니라, 선택적 물질수송, 신호전달등 중요한 기능을 수행한다. 원형질막의 세포질쪽 지질층은 주로 Phosphatidylethanolamine (PE), Phosphatidylserine (PS), Phosphatidylinositides (PIs) 등의 인지질로 구성되어 있는데, 다양한 단백질들이 이들과 직접 결합하거나 이들의 성질을 이용해 원형질막으로 위치된다. 본 연구에서는 원형질막의 안쪽 지질층에 존재하는 특정 인지질과 결합하여, 원형질막에 위치되는 단백질을 과발현시켰을 때 나타나는 세포의 모양변화를 분석하였다. 이를 위해 PS와 PI등과의 선택적 결합으로 원형질막에 위치하는 단백질들과 소수성 또는 정전기적 상호작용으로 원형질막으로 위치되는 단백질들을 HEK293T세포에 과발현시켜 보았다. 그 결과, 대조군으로 사용한 EGFP 단백질과 PI4P에 선택적으로 결합하는 EGFP-P4M-SidM단백질의 발현은 세포의 모양변화를 유도하지 않았다. 반면, PI(4,5)P₂에 결합하는 EGFP-PLCδ1(PH), PI(3,4,5)P₃에 결합하는 EGFP-AKT1(PH), PI4P와 PI(4,5)P₂에 결합하는 OSH2(PH)x2-EGFP의 발현은 세포의 크기가 줄어드는 수축현상이 일어나면서, Lamilapodia나 Filopodia가 형성되는 것을 확인할 수 있었다. 반면에, PS결합을 통해 원형질막에 위치되는 Lact-C2-EGFP과 소수성결합에 의해 원형질막에 위치되는 ApPDE4 long-form인 L(N20)-EGFP이나, 정전기적인 결합을 통해 원형질막으로 위치되는 ApPDE4 short-form인 S(N-UCR1-2)-EGFP단백질의 경우는 세포의 크기가 줄어드는 수축현상은 일어나지만 Lamilapodia나 Filopodia와 같은 모양변화는 나타나지 않는 것을 확인할 수 있었다. 본 연구를 통해, 특정 인지질에 결합하여 원형질막에 위치되는 단백질들이 각각의 인지질 결합 특성에 따라 분류 가능한 세포의 모양변화가 일어남을 확인할 수 있었다.

• Applied Biological Chemistry
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• 제34권2호
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• pp.117-124
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• 1991

기주 식물에 대한 인산공급이 근류의 인산 형태와 bacteroid의 인산흡수에 미치는 영향을 살펴보고자 B. japonicum MN 110을 접종한 대두식물을 1.0, 0.25, 0.05 mM-P 조건에서 재배하였다. 1.0, 0.25, 0.05 mM-P 조건에서 재배한 대두식물의 근류 Pi 농도는 각각 4.1, 2.5, 2.0 mM이었으며 인산공급이 근류의 Pi, SOP, lOP의 분포에는 유의성 있는 영향을 미치지 못하였다. 0.25, 0.05 mM-P 조건에서 재배한 대두식물의 유엽 Pi 농도는 대조구(1.0 mM-P)의 각각 33%, 20%이었으며 노엽 Pi 농도는 대조구의 각각 16%, 7%였다. 잎에서는 인산 결핍이 전인산에 대한 Pi의 분포를 감소시켰으나 lOP의 분포는 증가시켰다. 근류내의 bacteroid의 수는 $0.87-1.30{\times}10^{11}\;/GFW\;nodule$이었으며, 인산흡수도는 15-16 pmoles /min/$10^8$ bacteroid로 기주식물의 인산공급에는 영향을 받지 않았다 이러한 결과는 인산결핍 대두식물의 근류내 Pi농도는 bacteroid의 생육과 번식에 충분하며 인산결핍에 의하여 기주식물의 생육이 저해될 때에도 bacteroid의 인산흡수는 억제상태에 있음을 나타낸다.