AKT가 SV-HUC-1 세포주의 형질변화에 미치는 영향

김민규(Min Kyu Kim),이병란(Byung Lan Lee) 대한해부학회 2007 Anatomy & Cell Biology Vol.40 No.4

세포신호전달 물질인 protein kinase B (PKB/AKT)는 여러 종류의 세포에서 세포의 증식을 증가시키거나 세포사멸의 억제를 통해 발암을 일으키는 것으로 알려져 있지만 그 반대의 결과도 보고되고 있고, 또한 그 작용기전은 세포의 종류에 따라 다양하다. 지금까지 정상 방광상피세포에서 AKT의 기능은 잘 알려져 있지 않으므로 본 연구는 사람 이행상피세포주인 SV-HUC-1에서 AKT 활성을 억제시킨 후, AKT가 세포성장에 미치는 영향을 관찰하고 그 조절 기전을 연구하였다. 그 결과 유전자 이입을 통한 dominent negative AKT (DN-AKT)의 과발현으로 유도한 AKT의 활성억제가 crystal violet 염색과 flow cytometry에서 세포 성장률 감소와 G1 phase 세포들의 비율 증가를 초래함이 관찰되었다. 또한, 이때 p27kip1과 전사인자인 forkhead transcription factor (FKHR)의 단백발현이 증가함이 관찰되었다. 위 결과로부터 AKT의 활성은 방광상피세포의 세포성장에 기여하며 이때 세포주기 조절 단백인 p27kip1과 FKHR가 중요한 역할을 수행하는 것으로 사료되었다. Although protein kinase B (PKB, AKT) has been investigated extensively for its roles in oncogenic transformation and apoptotic prevention, controversial results are also reported. Here we assessed the role of AKT in the cell growth and expression of a key set of cell cycle regulators in the human normal uroepithelial cell line, SVHUC-1. AKT activity was suppressed by permanent transfection of dominent negative (DN)-AKT with Lipofectamine Plus. Cell viability was measured by the crystal violet assay. DNA contents stained by propidium iodide were measured by flow-cytometry for cell cycle analysis. Cell growth curve showed that overexpression of DN-AKT which suppressed the AKT activity decreased the cell growth. In the cell cycle analysis, overexpression of DN-AKT resulted in a 6% increase in the proportion of cells in G1 phase. Furthermore, DN-AKT overexpression increased the p27Kip1 protein expression and the activation of a transcription factor FKHR, which induces p27Kip1 transcription. Our results suggest that, in normal uroepithelial cells, AKT activation increases the cell growth through the influence on p27Kip1 expression and FKHR activation. Thus, AKT may be used as a biomarker for tumor transformation of bladder uroepithelial cells.

Connexin43 and zonula occludens-1 are targets of Akt in cardiomyocytes that correlate with cardiac contractile dysfunction in Akt deficient hearts

Ock, Sangmi,Lee, Wang Soo,Kim, Hyun Min,Park, Kyu-Sang,Kim, Young-Kook,Kook, Hyun,Park, Woo Jin,Lee, Tae Jin,Abel, E.D.,Kim, Jaetaek Elsevier 2018 Biochimica et biophysica acta Vol.1864 No.4

<P><B>Abstract</B></P> <P>While deletion of <I>Akt1</I> results in a smaller heart size and <I>Akt2</I> <SUP>−/−</SUP> mice are mildly insulin resistant, <I>Akt1</I> <SUP>−/−</SUP>/<I>Akt2</I> <SUP>−/−</SUP> mice exhibit perinatal lethality, indicating a large degree of functional overlap between the isoforms of the serine/threonine kinase Akt. The present study aimed to determine the cooperative contribution of Akt1 and Akt2 on the structure and contractile function of adult hearts. To generate an inducible, cardiomyocyte-restricted Akt2 knockout (KO) model, <I>Akt2</I> <SUP>flox/flox</SUP> mice were crossed with tamoxifen-inducible MerCreMer transgenic (MCM) mice and germline <I>Akt1</I> <SUP>−/−</SUP> mice to generate the following genotypes:<I>Akt1</I> <SUP>+/+</SUP>; <I>Akt2</I> <SUP>flox/flox</SUP> (WT), <I>Akt2</I> <SUP>flox/flox</SUP>; α-MHC-MCM (<I>iAkt2</I> KO), <I>Akt1</I> <SUP>−/−</SUP>, and <I>Akt1</I> <SUP>−/−</SUP>; <I>Akt2</I> <SUP>flox/flox</SUP>; α-MHC-MCM mice (<I>Akt1</I> <SUP>−/−</SUP>/<I>iAkt2</I> KO). At 28 days after the first tamoxifen injection, <I>Akt1</I> <SUP>−/−</SUP>/<I>iAkt2</I> KO mice developed contractile dysfunction paralleling increased atrial and brain natriuretic peptide (ANP and BNP) levels, and repressed mitochondrial gene expression. Neither cardiac fibrosis nor apoptosis were detected in <I>Akt1</I> <SUP>−/−</SUP>/<I>iAkt2</I> KO hearts. To explore potential molecular mechanisms for contractile dysfunction, we investigated myocardial microstructure before the onset of heart failure. At 3 days after the first tamoxifen injection, <I>Akt1</I> <SUP>−/−</SUP>/<I>iAkt2</I> KO hearts showed decreased expression of connexin43 (Cx43) and connexin-interacting protein zonula occludens-1 (ZO-1). Furthermore, Akt1/2 silencing significantly decreased both Cx43 and ZO-1 expression in cultured neonatal rat cardiomyocytes in concert with reduced beating frequency. Akt1 and Akt2 are required to maintain cardiac contraction. Loss of Akt signaling disrupts gap junction protein, which might precipitate early contractile dysfunction prior to heart failure in the absence of myocardial remodeling, such as hypertrophy, fibrosis, or cell death.</P> <P><B>Highlights</B></P> <P> <UL> <LI> Cardiac <I>Akt1</I> <SUP>-/-</SUP>/<I>iAkt2</I> KO in mice reduces cardiac size and induces contractile dysfunction without myocardial remodeling. </LI> <LI> <I>Akt1</I> <SUP>-/-</SUP>/<I>iAkt2</I> KO hearts revealed decreased expression of Cx43 and connexin-interacting protein ZO-1. </LI> <LI> Akt isoforms may play important roles to maintain cardiac contractile function through gap junction protein stability. </LI> </UL> </P>

Discovery and Evaluation of Polymorphisms in the AKT2 and AKT3 Promoter Regions for Risk of Korean Lung Cancer

Sung, Jae-Sook,Park, Kyong-Hwa,Kim, Seung-Tae,Kim, Yeul-Hong Korea Genome Organization 2012 Genomics & informatics Vol.10 No.3

AKT is a signal transduction protein that plays a central role in the tumorigenesis. There are 3 mammalian isoforms of this serine/threonine protein kinase-AKT1, AKT2, and AKT3-showing a broad tissue distribution. We first discovered 2 novel polymorphisms (AKT2 -9826 C/G and AKT3 -811 A/G), and we confirmed 6 known polymorphisms (AKT2 -9473 C/T, AKT2 -9151 C/T, AKT2 -9025 C/T, AKT2 -8618G/A, AKT3 -675 A/-, and AKT3 -244 C/T) of the AKT2 and AKT3 promoter region in 24 blood samples of Korean lung cancer patients using direct sequencing. To evaluate the role of AKT2 and AKT3 polymorphisms in the risk of Korean lung cancer, genotypes of the AKT2 and AKT3 polymorphisms (AKT2 -9826 C/G, AKT2 -9473 C/T, AKT2 -9151 C/T, AKT2 -9025 C/T, AKT2 -8618G/A, and AKT3 -675 A/-) were determined in 360 lung cancer patients and 360 normal controls. Statistical analyses revealed that the genotypes and haplotypes in the AKT2 and AKT3 promoter regions were not significantly associated with the risk of lung cancer in the Korean population. These results suggest that polymorphisms of the AKT2 and AKT3 promoter regions do not contribute to the genetic susceptibility to lung cancer in the Korean population.

Akt3 knockdown induces mitochondrial dysfunction in human cancer cells

Kim, Minjee,Kim, Young Yeon,Jee, Hye Jin,Bae, Sun Sik,Jeong, Na Young,Um, Jee-Hyun,Yun, Jeanho Oxford University Press 2016 Acta biochimica et biophysica Sinica Vol.48 No.5

<P>Akt/PKB plays a pivotal role in cell proliferation and survival. However, the isotype-specific roles of Akt in mitochondrial function have not been fully addressed. In this study, we explored the role of Akt in mitochondrial function after stable knockdown of the Akt isoforms in EJ human bladder cancer cells. We found that the mitochondrial mass was significantly increased in the Akt1-and Akt3-knockdown cells, and this increase was accompanied by an increase in TFAM and NRF1. Akt2 knockdown did not cause a similar effect. Interestingly, Akt3 knockdown also led to severe structural defects in the mitochondria, an increase in doxorubicin-induced senescence, and impairment of cell proliferation in galactose medium. Consistent with these observations, the mitochondrial oxygen consumption rate was significantly reduced in the Akt3-knockdown cells. An Akt3 deficiency-induced decrease in mitochondrial respiration was also observed in A549 lung cancer cells. Collectively, these results suggest that the Akt isoforms play distinct roles in mitochondrial function and that Akt3 is critical for proper mitochondrial respiration in human cancer cells.</P>

Direct tyrosine phosphorylation of Akt/PKB by epidermal growth factor receptor

배순식,최장현,윤성지,김은경,오용석,김치대,서판길,Bae, Sun-Sik,Choi, Jang-Hyun,Yun, Sung-Ji,Kim, Eun-Kyung,Oh, Yong-Suk,Kim, Chi-Dae,Suh, Pann-Ghill Korean Society of Life Science 2007 생명과학회지 Vol.17 No.2

Akt/PKB는 세포의 증식, 분화, 사멸, 혈관신생 등 매우 많은 생리활성 조절에 있어 매우 중요한 역할을 수행한다. 우리는 Akt/PKB의 tyrosine잔기의 인산화가 $Thr^{\308}$ 인산화에 필수적임을 밝혔다. COS-7 세포주에 EGF를 처 리하면 Akt/PKB의 tyrosine 잔기에 인산화가 촉진되었으며 이러한 인산화 촉진은 Akt/PKB에 myristoylation site를 이용해 세포막으로 이동시키면 더욱 더 증가하였다. 특히, 분리된 Akt/PKB와 EGF 수용체를 이용해 인산화 반응을 실시하면 tyrosine잔기의 인산화뿐만 아니라 $Ser^{\473}$에 대한 인산화도 증가하였다. 더욱이 tyrosine잔기에 인산화 된 Akt/PKB는 활성화된 EGF 수용체와 직접적인 결합을 이루고 있음을 확인하였다. 마지막으로 예측되는 tyrosine 잔기인 $(Tyr^{\326})$을 Alanine으로 치환하면 정상 Akt/PKB뿐만 아니라 활성화된 Akt/PKB의 EGF에 의한 $Thr^{\308}$ 인산화가 사라짐을 확인하였다. 이러한 결과들을 바탕으로 EGF 수용체에 의한 직접적인 Akt/PKB의 tyrosine 인산화는 EGF에 의한 많은 생리활성 조절기전의 또 다른 기전이라 볼 수 있다. Akt/PKB plays pivotal roles in many physiological responses such as proliferation, differentiation, apoptosis, and angiogenesis. Here we show that tyrosine phosphorylation of Akt/PKB is essential for the subsequent phosphorylation at $Thr^{\308}$. Tyrosine phosphorylation of Akt/PKB was induced by stimulation of COS-7 cells with epidermal growth factor receptor (EGF) and its phosphorylation was significantly enhanced by constitutive targeting of Akt/PKB to the plasma membrane by myristoylation. Interestingly, incubation of affinity purified Myc-tagged Akt/PKB with purified EGF receptor resulted in tyrosine phosphorylation as well as $Ser^{\473}$ phosphorylation of Akt/PKB. In addition, tyrosine-phosphorylated Akt/PKB could directly associate with activated EGF receptor in vitro. Finally, alanine mutation at putative tyrosine phosphorylation site $(Tyr^{\326})$ abolished EGF induced $Thr^{\308}$ phosphorylation of wild type as well as constitutively active form of Akt/PKB. Given these results we suggest here that direct tyrosine phosphorylation of Akt/PKB by EGF receptor could be another mechanism of EGF-induced control of many physiological responses.

HT-29 대장암세포에서 Akt 활성 저해에 따른 셀레늄의 세포 증식억제 효과

Song Yi Park(박송이),In-Seop Kim(김인섭),Se Hee Lee(이세희),Sol Hwa Lee(이솔화),Da Woon Jung(정다운),Ock Jin Park(박옥진),Young Min Kim(김영민) 한국생명과학회 2012 생명과학회지 Vol.22 No.1

Akt는 세포의 증식과 분화에 관여하며 많은 암종에서 과발현되어 있다는 것이 보고되었다. 본 연구에서는 Akt의 조절을 통한 셀레늄의 HT-29 세포의 세포증식억제 시너지효과를 확인하였다. 셀레늄을 농도별과 시간별로 처리하였을 때 HT-29 세포의 증식이 억제되었고, apoptosis가 일어남을 확인하였다. 셀레늄을 농도별로 처리하여 Western blotting 및 immunofluorescence를 실시한 결과 Akt의 인산화가 저해되었고 COX-2의 발현도 저해되었다. 또한 Akt 저해제인 LY294002를 처리한 결과, HT-29 대장암세포의 증식이 억제되었으며, LY294002를 셀레늄과 병행처리하였을 때 셀레늄에 의한 세포증식억제 효과가 더 강하게 나타나는 것을 확인하였다. Akt siRNA에 의한 Akt의 불활성화는 non-transfected 세포에 비하여 HT-29 세포의 성장을 더 강하게 억제하였으며, Akt가 불활성화 되었을 때 COX-2의 발현 역시 non-transfected 세포에 비하여 감소된 것을 확인하였다. 따라서 HT-29 세포에서 셀레늄의 세포증식억제 효과는 Akt와 COX-2 신호분자의 조절을 통해 일어나며, Akt 의 저해는 셀레늄의 대장암세포증식 억제에 시너지 효과를 나타냄을 확인하였다. Akt is known to play an important role in cell proliferation and differentiation, and is also over-expressed in several types of cancer cells. In this study, we explored the anti-proliferative effects of selenium in HT-29 colon cancer cells, mediated through effects on Akt and COX-2. Selenium treatments at different concentrations and for different durations inhibited proliferation of HT-29 colon cancer cells and increased apoptotic cell death. Selenium treatment decreased Akt phosphorylation and COX-2 expression. Treatment with LY294002 (an Akt inhibitor) decreased proliferation of HT-29 cells, while a combined treatment with LY294002 and selenium resulted in even further decreases in cell proliferation. Inactivation of Akt by Akt siRNA treatment abolished these inhibitory effects on cell growth. COX-2 expression decreased in Akt transfected cells compared to non-transfected cells. These results suggest that selenium induced both anti-proliferative and apoptotic effects by inhibiting Akt phosphorylation and COX-2 expression. Selenium treatment also appeared to induce synergistic anti-proliferative effects by inhibition of Akt in HT-29 colon cancer cells.

Discovery and Evaluation of Polymorphisms in the AKT2 and AKT3 Promoter Regions for Risk of Korean Lung Cancer

성재숙,김열홍,박경화,김승태 한국유전체학회 2012 Genomics & informatics Vol.10 No.3

AKT is a signal transduction protein that plays a central role in the tumorigenesis. There are 3 mammalian isoforms of this serine/threonine protein kinase-AKT1, AKT2, and AKT3-showing a broad tissue distribution. We first discovered 2 novel polymorphisms (AKT2 -9826 C/G and AKT3 -811 A/G), and we confirmed 6 known polymorphisms (AKT2 -9473 C/T, AKT2 -9151 C/T,AKT2 -9025 C/T, AKT2 -8618G/A, AKT3 -675 A/-, and AKT3 -244 C/T) of the AKT2 and AKT3 promoter region in 24 blood samples of Korean lung cancer patients using direct sequencing. To evaluate the role of AKT2 and AKT3 polymorphisms in the risk of Korean lung cancer, genotypes of the AKT2 and AKT3 polymorphisms (AKT2 -9826 C/G, AKT2 -9473 C/T, AKT2 -9151 C/T, AKT2 -9025 C/T,AKT2 -8618G/A, and AKT3 -675 A/-) were determined in 360 lung cancer patients and 360 normal controls. Statistical analyses revealed that the genotypes and haplotypes in the AKT2 and AKT3 promoter regions were not significantly associated with the risk of lung cancer in the Korean population. These results suggest that polymorphisms of the AKT2 and AKT3 promoter regions do not contribute to the genetic susceptibility to lung cancer in the Korean population.

The Significance of p-AKT1 as a Prognostic Marker and Therapeutic Target in Patients With Hormone Receptor-Positive and Human Epidermal Growth Factor Receptor-2-Positive Early Breast Cancer

김지예,박찬섭,장세경,설혜실,성민기,노우철,박인철,김현아 한국유방암학회 2022 Journal of breast cancer Vol.25 No.5

Purpose: Phosphorylated AKT1 (p-AKT1) at Ser473 is a functional isoform of AKT and a key component of the PI3K/mTOR/AKT pathway. This study aimed to evaluate the prognostic significance of p-AKT1 (Ser473) based on the molecular subtypes of breast cancer. Methods: To investigate the prognostic value of p-AKT1 (Ser473), we performed a retrospective chart review of patients with breast cancer. Data on p-AKT1 (Ser473) positivity, hormone receptor (HR) status, human epidermal growth factor receptor 2 (HER2) expression status, and other clinicopathological factors were obtained. Furthermore, the therapeutic effect of blocking p-AKT1 (Ser473) in breast cancer cells was evaluated using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, cell apoptosis assay, apoptosis protein array, and western blot analysis. Results: A total of 3,044 patients were evaluated, and the median follow-up time was 43 (range: 0–125) months. In patients with HR-positive and HER2-positive disease, the p-AKT1 (Ser473)-positive group had worse disease-free survival (DFS) than the p-AKT1 (Ser473)-negative group (hazard ratio, 1.9; 95% confidence interval, 1.1–3.5; p = 0.024). In the multivariate analysis, p-AKT1 (Ser473) remained a significantly worse prognostic factor in patients with HR-positive/HER2-positive breast cancer (p = 0.03). There was no difference in DFS according to p-AKT1 (Ser473) status among patients with other breast cancer subgroups. In vitro analysis showed that blocking p-AKT1 (Ser473) levels enhanced trastuzumab-induced cell death in HR-positive/HER2-positive and p-AKT1 (Ser473)-positive breast cancer cells. Conclusion: p-AKT1 (Ser473) is a prognostic marker for poor outcomes in patients with HR-positive/HER2-positive breast cancer and may have a potential value as a therapeutic target.

Regulation of Skeletal Muscle Differentiation by Akt

Dae Han Woo(우대한),Sung Ji Yun(윤성지),Eun Kyoung Kim(김은경),Jung Min Ha(하정민),Hwa Kyoung Shin(신화경),Sun Sik Bae(배순식) 한국생명과학회 2012 생명과학회지 Vol.22 No.4

Akt는 다양한 세포에서 성장, 발달, 증식, 분화와 같은 생리적 활성에 중요한 역할을 하고 골격근 세포에서 Akt는 재생 및 비대와 위축을 조절한다고 알려져 있다. 골격근 세포의 분화에 있어서 Akt의 역할을 밝히고자 본 연구를 수행하였다. 골격근 세포를 분화 시키기 위해 고밀도 및 저농도의 serum 상태에서 배양하며, 분화된 C2C12 근아세포는 둥근 모양에서 다핵을 가진 긴 모양으로 바뀐다. 이러한 형태학적 변화는 분화 시킨 후 2일부터 일어났다. 또한, 골격근 분화 표지인자인 myogenin D와 myogenin G의 발현은 2일 후 관찰되었다. C2C12세포주에 Akt1 또는 Akt2의 발현을 저하시키면 이와 더불어 골격근으로의 분화도 저해됨을 확인하였고, 이와는 반대로 Akt1 또는 Akt2를 과발현 시키면 골격근으로 분화가 촉진됨을 알 수 있었다. 이와 더불어 Akt의 활성은 분화유도 2일 후부터 관찰되었고 7일 이후로는 감소하였다. Kruppel-like factor 4의 발현은 6일부터 증가하는 것이 관찰이 되었다. Kruppel-like factor 4의 발현 또한 Akt1 또는 Akt2의 발현양이 감소된 C2C12 근아세포에서 줄어들어 있는 것을 확인하였다. 또한 Kruppel-like factor 4의 프로모터 부위에 대한 전사조절능력이 Akt1 또는 Akt2의 발현을 저하시켰을 때 같이 떨어짐을 확인하였다. 이러한 결과들로 보아 Akt가 골격근 분화를 조절하는데 있어 중요하며, Kruppel-like factor 4 발현이 이를 조절하는 데 있어 중요한 역할을 할 것이라 판단된다. Akt plays an important role in a variety of cellular physiologies such as growth, proliferation, and differentiation. In skeletal muscle, Akt has been implicated in regulating regeneration, hypertrophy, and atrophy. In this study, the role of Akt has been examined during skeletal muscle differentiation. Culturing C2C12 myoblasts under low serum (1% horse serum) and high density converted cell morphology from a round shape to an elongated and multi-nucleated shape. Morphological changes were initiated from day 2 of differentiation. In addition, the expression of both myogenin G and myogenin D was elevated from day 2 of differentiation. Skeletal muscle differentiation was abolished by silencing Akt1 or Akt2, but was significantly enhanced by the over-expression of either Akt1 or Akt2. The activation of Akt was observed from day 2 of differentiation and disappeared after day 7. The expression of kruppel-like factor 4 was observed from day 6 of differentiation. Moreover, this expression was blocked in cells silencing either Akt1 or Akt2. In addition, the promoter activity of kruppel-like factor 4 was significantly reduced in cells silencing Akt1 or Akt2. These results suggest that Akt regulates skeletal muscle differentiation through the regulation of kruppel-like factor 4 expression.

저칼륨혈증 흰쥐 신장에서 Akt, p-Akt, ERK 및 p-ERK 단백발현의 변화

배춘상(Choon Sang Bae),조혜정(Hye Jung Cho),안규윤(Kyu Yoon Ahn) 대한체질인류학회 2017 해부·생물인류학 (Anat Biol Anthropol) Vol.30 No.3

저칼륨혈증은 신장의 형태학적 변화와 대사성 알칼리증을 유발시키는 것으로 알려져 있다. 이전 결과들은 저칼륨혈증의 병태생리학적 변화에 K+평형 조절 이온채널, pump 유전자, NF-E2-related factor 2 (Nrf2) 전사유전자를 포함한 다양한 유전자가 관여할 것이라는 가능성을 제시하였다. 이에 본 연구는 칼륨제한 식이 기간에 따른 흰쥐 신장 내 Akt와 p-Akt 및 ERK와 p-ERK의 발현 및 분포의 변화를 Western 분석과 면역조직화학 방법으로 관찰함으로써, 저칼륨혈증이 신장에서 AKT/ERK 인산화에 영향을 미치는지를 확인하였다. Western 분석소견에서 Akt 및 p-Akt 단백질 발현은 칼륨제한 식이가 진행될수록 증가하는 양상을 보였으며, ERK 및 p-ERK 단백질발현은 칼륨제한 식이 2주군에서 정상 식이군에 비해 약간 증가하는 양상을 보였다. 면역조직화학 소견에서 Akt 단백의 면역반응성은 먼쪽곱슬세관, 겉질곧은부분 및 속질곧은부분에서 중등도의 발현을 보였다. 칼륨제한 식이군의 Akt의 면역반응성은 칼륨제한 식이가 진행될수록 바깥속질집합관에서 현저히 증가하였다. 칼륨제한 식이군의 p-Akt의 면역반응성은 칼륨제한 식이 2주군의 먼쪽곱슬세관, 치밀반점과 속질곧은부분에서 증가하였고 토리쪽곱슬세관에서는 중등도의 발현을 보였다. 칼륨제한 식이군의 ERK의 면역반응성은 칼륨제한 식이 2주군과 3주군의 바깥속질집합관에서 현저히 증가하였으며 먼쪽곱슬세관, 겉질집합관에서는 증등도의 증가를 보였다. 칼륨제한 식이군의 p-ERK의 발현부위는 정상식이군과 비교해 차이가 없었으나 면역반응성은 칼륨제한 식이 2주군의 바깥속 질집합관의 핵에서 현저히 증가하였다. 이상의 결과 저칼륨혈증 시 p-Akt의 발현은 칼륨제한 식이가 길어질수록 점진적으로 증가하였지만, p-ERK의 발현은 칼륨제한 식이 2주군에서 현저히 증가되었다. 따라서 저칼륨 상태에서의 Akt 및 ERK 인산화의 촉진은 이온채널 및 이온수송체 유전자 조절뿐만 아니라 세포 내 신호전달에 있어 중요한 역할에 관여할 것임을 시사해 주었다. Hypokalemia causes metabolic alkalosis and morphological changes of the kidney. K+ balance is regulated not only by ion channels or pump gene, but also by various genes including NF-E2-related factor 2 (Nrf2). Previous study suggested the possibility that Akt and ERK kinase may be involved in Nrf2 transcriptional gene activation. In present study, we investigate the alterations of Akt, p-Akt, ERK, p-ERK protein in both normal kidney and K+-deficient diet kidney using Western blot analysis, and immunohistochemisrty. Our western blot data showed that the expression of Akt and p-Akt was increased gradually in K+-depleted diet (from 1W-3W) compared to normal group. The expression of ERK and p-ERK was markedly increased in K+-depleted diet 2W in comparison with normal group. Based on our immunostaining results, Akt protein immunoreactivity was prominently increased in outer medullary collecting duct, especially in K+-depleted diet 2 weeks. The localization of p-Akt proteins in K+-depleted groups was not different from normal group, but the immunoreactivity was significantly increased in distal convoluted tubule, macula densa and outer medullary thick ascending limb in K+-depleted diet 1 and 2 weeks groups. ERK protein immunoreactivity was prominently increased in outer medullary collecting duct, especially in K+-depleted diet 2 and 3 weeks. The localization of p-ERK proteins in K+-depleted groups was not different from normal group, but the immunoreactivity was prominently increased in the nucleus of outer medullary collecting duct especially in K+-depleted diet 2 weeks. Taken together, we suggest that the expression of p-Akt was gradually increased in K+-depleted groups of kidney, but the expression of p-ERK was markedly increased in K+-depleted diet 2 week group. Hence, the promotion of AKT and ERK phosphorylation in hypokalemic condition may be involved in the regulation of ion channels, ion transporters and subsequent intracellular signal transduction.