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Identifying the sources and weathering degree of sediments in the northern deserts of China is crucial to understand the evolution of the Asian monsoon system and corresponding induced changes in surface processes. Great progress has been made in the reconstruction of paleoclimate and paleoenvironment from the Early Holocene onwards based on chronostratigraphy and proxies of aeolian sedimentary sequences in the Mu Us Desert (MUD). However, previous studies have given rise to some controversial over the timing of the Holocene optimum period, and there is a scarcity of studies on the sediment provenance evolution of the desert during the Holocene and its internal relationship with the East Asian summer (EASM) and winter (EAWM) monsoons. Here, we provide new elemental geochemical data from a fluvial–aeolian sedimentary sequence in the southern MUD; we then combine these data with other available elemental data and hydroclimatology information to investigate the characteristics of sediment source evolution and their possible connection with the EASM and EAWM on the southern margin of the MUD-desert/loess transition zone since the latter part of the Early-Holocene. The elemental composition and geochemical characteristics of the studied sedimentary sequence differed markedly in the periods before and after 6.4 ka BP, suggesting a change in the source of the sediments across this boundary. From the late Early-Holocene (10.6 cal. ka BP), sediments were dominated by the chemical weathering products of local source material, with a gradual increase in chemical weathering over time, indicating a gradual strengthening of the EASM; the EASM reached its peak Holocene strength at 8.4–6.4 ka BP, accompanied by a moderate level of chemical weathering. After 6.4 ka BP, with the weakening of monsoon precipitation, the EAWM began to strengthen, and long-distance transport of source material from the west and northwest increased; however, chemical weathering of local source material driven by the EASM remained dominant until 4.0 ka BP. After 4.0 ka BP, the EAWM became dominant, and sediments showed strong sedimentary cycles, mainly transported from long-distance sources by aeolian activity, and the MUD became an important sedimentary sink for the northern-central deserts. After 0.7 ka BP, the sediments in the MUD became a mixture of different-age sediments due to river scouring and the transport of pre-existing sediments along the shoreline. Collectively, the elemental geochemical and other proxy data from Holocene sediments on the southern margin of the MUD record an inverse relationship between the EASM and EAWM; these two monsoonal circulations have alternated in their control of the sedimentary source characteristics and geomorphological environmental evolution processes across the southern margin of the MUD during the Holocene.

Abstract Because the number of clustering cores needs to be set before implementing the K-means algorithm, this type of algorithm often fails in applications with increasing data and changing distribution characteristics. This paper proposes an evolutionary algorithm DCC, which can dynamically adjust the number of clustering cores with data change. DCC algorithm uses the Gaussian function as the activation function of each core. Each clustering core can adjust its center vector and coverage based on the response to the input data and its memory state to better fit the sample clusters in the space. The DCC algorithm model can evolve from 0. After each new sample is added, the winning dynamic core can be adjusted or split by competitive learning, so that the number of clustering cores of the algorithm always maintains a better adaptation relationship with the existing data. Furthermore, because its clustering core can split, it can subdivide the densely distributed data clusters. Finally, detailed experimental results show that the evolutionary clustering algorithm DCC based on the dynamic core method has excellent clustering performance and strong robustness.

Abstract In this paper, we propose a structural developmental neural network to address the plasticity‐stability dilemma, computational inefficiency, and lack of prior knowledge in continual unsupervised learning. This model uses competitive learning rules and dynamic neurons with information saturation to achieve parameter adjustment and adaptive structure development. Dynamic neurons adjust the information saturation after winning the competition and use this parameter to modulate the neuron parameter adjustment and the division timing. By dividing to generate new neurons, the network not only keeps sensitive to novel features but also can subdivide classes learnt repeatedly. The dynamic neurons with information saturation and division mechanism can simulate the long short‐term memory of the human brain, which enables the network to continually learn new samples while maintaining the previous learning results. The parent‐child relationship between neurons arising from neuronal division enables the network to simulate the human cognitive process that gradually refines the perception of objects. By setting the clustering layer parameter, users can choose the desired degree of class subdivision. Experimental results on artificial and real‐world datasets demonstrate that the proposed model is feasible for unsupervised learning tasks in instance increment and class increment scenarios and outperforms prior structural developmental neural networks.

Abstract Systematic quantification of phosphoprotein within cell signaling networks in solid tissues remains challenging and precise quantification in large scale samples has great potential for biomarker identification and validation. We developed a reverse phase protein array (RPPA) based phosphor-antibody characterization approach by taking advantage of the lysis buffer compatible with alkaline phosphatase (AP) treatment that differs from the conventional RPPA antibody validation procedure and applied it onto fresh frozen (FF) and formalin-fixed and paraffin-embedded tissue (FFPE) to test its applicability. By screening 106 phospho-antibodies using RPPA, we demonstrated that AP treatment could serve as an independent factor to be adopted for rapid phospho-antibody selection. We also showed desirable reproducibility and specificity in clincical specimens indicating its potential for tissue-based phospho-protein profiling. Of further clinical significance, using the same approach, based on melanoma and lung cancer FFPE samples, we showed great interexperimental reproducibility and significant correlation with pathological markers in both tissues generating meaningful data that match clinical features. Our findings set a benchmark of an efficient workflow for phospho-antibody characterization that is compatible with high-plex clinical proteomics in precison oncology.

The cold regions of China (CRC) are important and vulnerable freshwater recharge areas on land, and any changes in them are related to the survival of millions of people in East Asia. However, for nearly half a century, in cold regions, the extreme temperature response to global warming is still poorly understood. In this study, we systematically studied the temperature extreme changes in cold regions of China since 1961 and discussed the possible circulation factors in detail. The results showed that 1) the warming magnitudes in cold nights and warm nights are greater than those in cold days and warm days, and decreases in cold nights and cold days and increases in warm days and warm nights appeared in almost all of cold regions of China. Most of the temperature indices displayed the largest magnitudes of warming in winter. 2) Spatially, for most of the temperature extremes, the stations located at Qinghai–Tibet Plateau (TPC) and Northwest China (NWC) showed a larger warming trend than that shown by the station at Northeast China (NEC). 3) The responses of temperature extremes at different cold regions to each circulation index are variable. Atlantic Multidecadal Oscillation (AMO) has a significant relationship with almost all the indices in cold regions of China. Almost all the temperature extremes of TPC and NWC showed closely relationship with the North Atlantic Oscillation (NAO), especially for diurnal temperature range (DTR), daily maximum temperature, and the cold extremes. Multivariate ENSO Index (MEI) is significantly related to most the temperature indices of Northwest China and Northeast China. However, MEI has a significant impact on only TPC’s diurnal temperature range and warmest night (TNx). 4) Atlantic Multidecadal Oscillation displayed significant relationships with most the temperature extremes in every season in cold regions of China. However, the summer and winter MEI and the summer and winter North Atlantic Oscillation showed significant impacts on only diurnal temperature range, daily minimum temperatures (TNm), and TNx.

Hallux valgus is a common form of foot deformity, and genetic factors contribute substantially to the pathogenesis of hallux valgus deformity. We conducted a genetic study on the structural variants underlying familial hallux valgus using whole exome sequencing approach. Twenty individuals from five hallux valgus families and two sporadic cases were included in this study. A total of 372 copy number variations were found and passed quality control filtering. Among them, 43 were only present in cases but not in controls or healthy individuals in the database of genomic variants. The genes covered by these copy number variations were enriched in gene sets related to immune signaling pathway, and cytochrome P450 metabolism. The hereditary CNVs demonstrate a dominant inheritance pattern. Two candidate pathogenic CNVs were further validated by quantitative-PCR. This study suggests that hallux valgus is a degenerative joint disease involving the dysregulation of immune and metabolism signaling pathways.

Abstract Alterations in DNAs could not reveal what happened in proteins. The accumulated alterations of DNAs would change the manifestation of proteins. Therefore, as is the case in cancer liquid biopsies, deep proteome profiling will likely provide invaluable and clinically relevant information in real-time throughout all stages of cancer progression. However, due to the great complexity of proteomes in liquid biopsy samples and the limitations of proteomic technologies compared to high-plex sequencing technologies, proteomic discoveries have yet lagged behind their counterpart, genomic technologies. Therefore, novel protein technologies are in urgent demand to fulfill the goals set out for biomarker discovery in cancer liquid biopsies. Notably, conventional and innovative technologies are being rapidly developed for proteomic analysis in cancer liquid biopsies. These advances have greatly facilitated early detection, diagnosis, prognosis, and monitoring of cancer evolution, adapted or adopted in response to therapeutic interventions. In this paper, we review the high-plex proteomics technologies that are capable of measuring at least hundreds of proteins simultaneously from liquid biopsy samples, ranging from traditional technologies based on mass spectrometry (MS) and antibody/antigen arrays to innovative technologies based on aptamer, proximity extension assay (PEA), and reverse phase protein arrays (RPPA).

Alzheimer’s disease (AD) is a common neurodegenerative disorder causing dementia in the elderly population. Functional disconnection of brain is considered to be the main cause of AD. In this study, we applied a newly developed association (Asso) mapping approach to directly quantify the functional disconnections and to explore the diagnostic effects for AD with resting-state functional magnetic resonance imaging data from 36 AD patients and 42 age-, gender-, and education-matched healthy controls (HC). We found that AD patients showed decreased Asso in left dorsoanterior insula (INS) while increased functional connections of INS with right medial prefrontal cortex (MPFC) and left posterior cingulate cortex (PCC). The changed Asso and functional connections were closely associated with cognitive performances. In addition, the reduced Asso and increased functional connections could serve as effective neuromarkers to distinguish AD patients from HC. Our research provided new evidence for functional disconnections in AD and demonstrated that functional disconnections between cognition-memory networks may be potential early biomarkers for AD.

Subependymal heterotopia (SEH) is a rare neuronal migration disorder consisting of gray matter nodules along the lateral ventricular walls and is often associated with other brain malformations. Despite most SEH cases showing epilepsy during their lifetimes, very few patients with asymptomatically familial SEH tend to cause misdiagnosis or missed diagnosis. We present four familial SEH cases without any positive symptoms and medical history, including two fetuses, who were diagnosed by MRI and confirmed by genetic testing with mutation of filamin A. This report emphasizes the role of MRI in the recognition of SEH at an early age of gestation and in asymptomatically familial SEH. MRI provides a fast, repeatable, reliable, and cheap choice for detecting and screening familial SEH.

The aim of this study was to explore the role of hsa_circRNA_0000205 (circ_0000205) in chondrocyte injury in osteoarthritis (OA) and the underlying mechanism. Expression of circ_0000205, microRNA (miR)-766-3p and a disintegrin and metalloproteinase with thrombospondin motif (ADAMTS)-5 was detected by quantitative real time (qRT)-polymerase chain reaction (PCR) and Western blot assays. Cell proliferation, apoptosis, and extracellular matrix (ECM) synthesis were examined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and 5-ethynyl-2-deoxyuridine assays, flow cytometry, and qRT-PCR and Western blot assays. The target relationship between miR-766-3p and circ_0000205 or ADAMTS5 was confirmed by luciferase reporter assay and RNA immunoprecipitation. IL-1β treatment could attenuate cell viability of primary chondrocytes and proliferating cell nuclear antigen (PCNA) and collagen II type alpha-1 (COL2A1) levels, and elevate apoptosis rate and cleaved caspase-3, ADAMTS5 and matrix metalloproteinase-13 (MMP13) levels, suggesting that IL-1β induced chondrocyte apoptosis and ECM degradation. Expression of circ_0000205 was up-regulated in OA tissues and IL-1β-induced primary chondrocytes, accompanied with miR-766-3p down-regulation and ADAMTS5 up-regulation. Knockdown of circ_0000205 could mitigate IL-1β-induced above effects and improve cell proliferation. Moreover, both depleting miR-766-3p and promoting ADAMTS5 could partially counteract circ_0000205 knockdown roles in IL-1β-cultured primary chondrocytes. Notably, circ_0000205 was verified as a sponge for miR-766-3p via targeting, and ADAMTS5 was a direct target for miR-766-3p. Silencing circ_0000205 could protect chondrocytes from IL-1β-induced proliferation reduction, apoptosis, and ECM degradation by targeting miR-766-3p/ADAMTS5 axis.

An excavated profile of aeolian-palaeosol-lacustrine sediments (the Wapianliang profile), located at the southeastern part of the Mu Us Desert, Northern China, was studied to reconstruct regional Holocene environmental changes. A chronology was established based on three AMS 14C and two OSL dates, and variations in the lithology and grain size, magnetic susceptibility, soil micromorphology, and chemical elements were used to explore the regional depositional environments during the Holocene. The results showed that since around 14 ka BP, this region had experienced seven alternations of wetting and drying. A shallow lake, which was identified by celadon lacustrine sediments with sporadic freshwater gastropod fossils, occurred in this area from around 13.0 ka BP to 9.9 ka BP. There existed two obvious intervals of soil formation, inferred from the environmental proxies of the palaeosol/sandy palaeosol layers, with relatively fine average grain-size, high magnetic susceptibility value, remarkable pedogenesis features, and strong chemical weathering, in particular, a well-developed palaeosol layer dating from the middle Holocene (8.6 ka BP to 4.2 ka BP). A weakly-developed palaeosol layer (from around 1.2 ka BP) at the upper part of the profile is possibly an indication of the Medieval Warm Period. This implies a forest steppe environment at both of these sedimentary stages. After 0.9 ka BP, a desert environment returned, analogous to before around 13.0 ± 1.4 ka BP, between 9.9 ± 1.1 ka BP to 8.6 ka BP, and between 4.2 ka BP to 1.6 ka BP, indicating the aggravation of aeolian activity and the expansion of mobile sand dunes. The variations in sedimentary environments were mainly triggered by changes in the East Asian Summer Monsoon (EASM).

Advances in immunotherapy have made an unprecedented leap in treating colorectal cancer (CRC). However, more effective therapeutic regimes need a deeper understanding of molecular architectures for precise patient stratification and therapeutic improvement. We profiled patients receiving neoadjuvant chemotherapy alone or in combination with immunotherapy (PD-1 checkpoint inhibitor) using Digital Spatial Profiler (DSP), a high-plex spatial proteogenomic technology. Compartmentalization-based high-plex profiling of both proteins and mRNAs revealed pronounced immune infiltration at tumor regions associated with immunotherapy treatment. The protein and the corresponding mRNA levels within the same selected regions of those patient samples correlate, indicating an overall concordance between the transcriptional and translational levels. An elevated expression of PD-L1 at both protein and the mRNA levels was discovered in the tumor compartment of immunotherapy-treated patients compared with chemo-treated patients, indicating potential prognostic biomarkers are explorable in a spatial manner at the local tumor microenvironment (TME). An elevated expression of PD-L1 was verified by immunohistochemistry. Other compartment-specific biomarkers were also differentially expressed between the tumor and stromal regions, indicating a dynamic interplay that can potentiate novel biomarker discovery from the TME perspectives. Simultaneously, a high-plex spatial profiling of protein and mRNA in the tumor microenvironment of colorectal cancer was performed.

Abstract Background Aberrant AKT activation is prevalent across human cancer lineages, providing an important therapeutic target. AKT comprises three isoforms that mediate critical non-redundant, even opposing functions in cancer pathophysiology. Therefore, targeting specific AKT isoforms in particular cancers may be more effective than pan-AKT inhibition while avoiding disadvantages of pan-AKT inhibition. Currently, AKT isoform-specific expression and activation in cancer are not clearly characterized. Methods We systematically characterized AKT isoform-specific expression and activation in 211 cancer cell lines derived from different lineages and genetic backgrounds using a reverse-phase protein array platform. Results We found that phosphorylation, but not expression, of AKT1 and AKT2 was coordinated in most but not all cells. Different cancer lineages displayed differential AKT1 and AKT2 expression and phosphorylation. A PIK3CA hotspot mutation H1047R but not E545K was associated with selective activation of AKT2 but not AKT1. Conclusions Our study identified and validated AKT isoform-specific expression and phosphorylation in certain cell lines and demonstrated that genetic changes can affect AKT isoform-specific activation. These results provide a more precise understanding of AKT isoform-specific signaling and, in addition, facilitate AKT isoform targeting for personalized cancer therapies.

Abstract Background Extracellular matrix (ECM) is a mediator of tumor progression. However, whether the alterations of the intraperitoneal ECM prior to tumor establishment affects the malignant progression of ovarian cancer remains elusive. Methods Apolipoprotein (ApoE) knock-out mice was used to analyze the intraperitoneal ECM alterations by quantification of the major components of ECM. ID8 cells were implanted in vivo to generate allografts and human ovarian cancer cell lines were characterized in vitro to assess the effects of ECM alterations on the malignant progression of ovarian cancer. Adhesion assay, immunochemistry, cytokines profile, proliferation assay, transwell invasion assay and western blot were used to determine the malignant phenotype of ovarian cancer cells. Results ApoE loss induced increased ECM deposition, which stimulated the adhesions of ovarian cancer cells. The adhesion-mediated focal adhesion kinase (FAK) signaling enhanced the invasive behaviors of ovarian cancer cells through activation of a ERK-MMP linkage. This ECM-induced signaling cascade was further confirmed in human ovarian cancer cell lines in vitro. Furthermore, reversal of the ECM accumulation with BAPN or abrogation of adhesion-induced ERK activation in ovarian cancer cells with MEK inhibitors (MEKi) was found to effectively delay ovarian cancer progression. Conclusions These findings identify the FAK-ERK activation in cell/matrix adhesion in the malignant progression of ovarian cancer and the efficiency of BAPN or MEKi for tumor suppression, providing an impetus for further studies to explore the possibility of new anticancer therapeutic combinations.

Abstract Background In 2016, it is estimated that there will be 62,700 new cases of kidney cancer in the United States, and 14,240 patients will die from the disease. Because the incidence of kidney renal clear cell carcinoma (KIRC), the most common type of kidney cancer, is expected to continue to increase in the US, there is an urgent need to find effective diagnostic biomarkers for KIRC that could help earlier detection of and customized treatment strategies for the disease. Accordingly, in this study we systematically investigated KIRC’s prognostic biomarkers for survival using the reverse phase protein array (RPPA) data and the high throughput sequencing data from The Cancer Genome Atlas (TCGA). Results With comprehensive data available in TCGA, we systematically screened protein expression based survival biomarkers in 10 major cancer types, among which KIRC presented many protein prognostic biomarkers of survival time. This is in agreement with a previous report that expression level changes (mRNAs, microRNA and protein) may have a better performance for prognosis of KIRC. In this study, we also identified 52 prognostic genes for KIRC, many of which are involved in cell-cycle and cancer signaling, as well as 15 tumor-stage-specific prognostic biomarkers. Notably, we found fewer prognostic biomarkers for early-stage than for late-stage KIRC. Four biomarkers (the RPPA protein IDs: FASN, ACC1, Cyclin_B1 and Rad51) were found to be prognostic for survival based on both protein and mRNA expression data. Conclusions Through pan-cancer screening, we found that many protein biomarkers were prognostic for patients’ survival in KIRC. Stage-specific survival biomarkers in KIRC were also identified. Our study indicated that these protein biomarkers might have potential clinical value in terms of predicting survival in KIRC patients and developing individualized treatment strategies. Importantly, we found many biomarkers in KIRC at both the mRNA expression level and the protein expression level. These biomarkers shared a significant overlap, indicating that they were technically replicable.

Stereo matching is one of the most important and challenging subjects in the field of stereo vision. The disparity obtained in stereo matching can represent depth information in 3-D world to a great extent and shows great importance in stereo field. In general, stereo-matching methods primarily emphasize static image. However, the information provided by dynamic scene can be used fully and effectively to improve the results of stereo matching for dynamic scene, such as video sequences. In this paper, we propose a dynamic scene-based local stereo-matching algorithm which integrates a cost filter with motion flow of dynamic video sequences. In contrast to the existing local approaches, our algorithm puts forward a new computing model which fully considers motion information in dynamic video sequences and adds motion flow to calculate suitable support weight for accurately estimating disparity. Our algorithm can perform as an edge-preserving smoothing operator and shows improved behavior near the moving edges. The experimental results show that the proposed method achieves a better depth map and outperforms other local stereo-matching methods in disparity evaluation.

With the intensification of global warming, spatiotemporal variations in the climate and their mechanisms have received increasing attention. Currently, the relationship between regional precipitation regime, large-scale circulation, and topography, particularly in high-altitude areas such as the Qinghai–Tibet Plateau, are not well understood. Herein, the spatial and temporal variability in the annual and intra-annual (wet and dry periods) precipitation at 33 stations in the Three-River Headwaters (TRH) region from 1967 to 2016 are analysed. Moreover, the empirical orthogonal function and wavelet transform coherence methods are used to analyse the relationships between the different modes of precipitation change and 14 atmospheric circulation indices. The following results were obtained. (1) The mean annual precipitation and mean dry period precipitation significantly increased over the studied period. Annual and intra-annual precipitation showed a spatial southeast-to-northwest decreasing trend. (2) Two main patterns of precipitation were observed during the studied period: a dominant pattern with high- and low-value centres located in southeast and northwest TRH, respectively, and a dipole pattern with more precipitation over southwest TRH and less precipitation over northeast TRH. (3) Precipitation had a negative correlation with latitude, positive correlation with longitude, and nonlinear relationship with elevation. (4) Precipitation changes over various parts of the studied domain were determined based on changes in the weather systems affecting the area, with different indices being correlated with different components during different times of the year.

In the original publication of this manuscript [1], Fig. 5E lower panel was incorrect due to an error in the preparation of these figures for publication. It was noticed that in the lower panel of Fig. 5E, one mouse image of ApoE−/− + PBS group (upper) was a photograph coming from ApoE−/− + BAPN pre-treatment group (lower). The corrected figure appears below. We apologize for any confusion this may have caused.

Extreme precipitation events affect the ecological environment and are also important for the sustainable development of regional socioeconomics. Although there are some local studies on extreme precipitation events in which the temporal and spatial variation characteristics of extreme precipitation events in non-monsoon regions (NMRs) are systematically assessed, detailed study on the driving mechanisms of variation are becoming increasingly important. In this study, nine extreme precipitation indices were used to analyze the characteristics of extreme precipitation event spatiotemporal variations in NMRs in China during 1961–2017. The results show that except for the consecutive dry days, which shows a significant decreasing trend ( P < 0.01) of −2.33 days/decade, all other indices showed obvious increasing trends, especially the indices of wet day precipitation (PRCPTOT), highest 5 day precipitation (RX5day) and light rain days (R5 mm), with significantly increasing trends ( P < 0.01) of 6.80 mm/decade, 0.73 mm/decade and 0.45 days/decade, respectively. In addition, a correlation analysis between altitude, longitude, latitude and extreme precipitation shows that stations at an altitude of more than 3500 m have significant correlations with both extreme precipitation and longitude in NMRs ( P < 0.05). In addition, results also indicated that there are significant relationships between extreme precipitation events in NMRs and large-scale ocean-atmosphere circulation patterns ( P < 0.05). The rapid increase in extreme precipitation indices over the past 20 years is closely related to the Atlantic Multidecadal Oscillation shift to a warm phase, while the Pacific Decadal Oscillation, El Niño–Southern Oscillation and Summer Monsoon Index show significant correlation with the extreme indices only in certain seasons ( P < 0.05).

Nanofluidic proteomic immunoassay (NIA), consisting of isoelectric focusing followed by sensitive chemiluminescence detection, has the potential to quantitatively characterize protein post-translational modifications that cause shifts in isoelectric point (pI). This protocol details the NIA analysis of protein phosphorylation using AKT as an example. This protocol can be used for two platforms, NanoPro 1000 and Peggy Sue, from ProteinSimple Company. NanoPro 1000 separates proteins based on charge. Peggy Sue separates proteins based on either charge or size. The platforms can analyze up to 96 samples at a time with lysates from as few as 25 cells per assay. Detailed information of the platforms is available on ProteinSimple’s website (http://www.proteinsimple.com).

The enzyme 5α-reductase, which converts testosterone to dihydrotestosterone (DHT), performs key functions in the androgen receptor (AR) signaling pathway. The three isoenzymes of 5α-reductase identified to date are encoded by different genes: SRD5A1, SRD5A2, and SRD5A3. In this study, we investigated mechanisms underlying androgen regulation of 5α-reductase isoenzyme expression in human prostate cells. We found that androgen regulates the mRNA level of 5α-reductase isoenzymes in a cell type-specific manner, that such regulation occurs at the transcriptional level, and that AR is necessary for this regulation. In addition, our results suggest that AR is recruited to a negative androgen response element (nARE) on the promoter of SRD5A3 in vivo and directly binds to the nARE in vitro. The different expression levels of 5α-reductase isoenzymes may confer response or resistance to 5α-reductase inhibitors and thus may have importance in prostate cancer prevention.

Frequent activation of the AKT serine-threonine kinase in cancer confers resistance to therapy. AKT is activated by a multi-step process involving phosphatidylinositide (PtdIns) phosphate-mediated recruitment of AKT and its upstream kinases, including 3-Phosphoinositide-dependent kinase 1 (PDK1), to the inner surface of the cell membrane. PDK1 in the appropriate context phosphorylates AKT at threonine 308 (T308) to activate AKT. Whether PtdIns(3,4,5)Ps (PtdInsP3) binding and AKT membrane translocation mediate functions other than formation of a functional PDK1::AKT complex have not been fully elucidated. We fused complementary fragments of intensely fluorescent protein (IFP) to AKT1 and PDK1 to induce a stable complex to study the prerequisites of AKT1 phosphorylation and function. In the stabilized PDK1-IFPC::IFPN-AKT1 complex, AKT1 T308 phosphorylation was independent of PtdIns, as demonstrated by treatment with Phosphatidylinositol 3 Kinase (PI3K) inhibitors. Further when interaction with PtdIns and the cell membrane was prevented by creating PH-domain mutants of AKT1 (R25A) and PDK1 (R474A), AKT1 phosphorylation on T308 was maintained in the PDK1-IFPC::IFPN-AKT1 complex. The PDK1-IFPC::IFPN-AKT1 complex was sufficient for phosphorylation of known AKT substrates, and conferred resistance to inhibitors of PI3K (LY294002, PI103, GDC0941 and TGX286) but not inhibitors of the downstream TORC1 complex (rapamycin). Thus the locus of action of targeted therapeutics can be elucidated by the constitutively active AKT1 complex. Our data indicate that PtdIns and membrane localization are not required for AKT phosphorylation and activation, but rather serve to induce a functional physical interaction between PDK1 and AKT. The PDK1-IFPC::IFPN-AKT1 complex provides a cell-based platform to examine specificity of drugs targeting PI3K pathway components.