Direct observations of energy transfer from resonant electrons to whistler-mode waves in magnetosheath of Earth.

Electromagnetic whistler-mode waves in space plasmas play critical roles in collisionless energy transfer between the electrons and the electromagnetic field. Although resonant interactions have been considered as the likely generation process of the waves, observational identification has been extremely difficult due to the short time scale of resonant electron dynamics. Here we show strong nongyrotropy, which rotate with the wave, of cyclotron resonant electrons as direct evidence for the locally ongoing secular energy transfer from the resonant electrons to the whistler-mode waves using ultra-high temporal resolution data obtained by NASA's Magnetospheric Multiscale (MMS) mission in the magnetosheath. The nongyrotropic electrons carry a resonant current, which is the energy source of the wave as predicted by the nonlinear wave growth theory. This result proves the nonlinear wave growth theory, and furthermore demonstrates that the degree of nongyrotropy, which cannot be predicted even by that nonlinear theory, can be studied by observations.

Collaborative Research Activities of the Arase and Van Allen Probes.

This paper presents the highlights of joint observations of the inner magnetosphere by the Arase spacecraft, the Van Allen Probes spacecraft, and ground-based experiments integrated into spacecraft programs. The concurrent operation of the two missions in 2017-2019 facilitated the separation of the spatial and temporal structures of dynamic phenomena occurring in the inner magnetosphere. Because the orbital inclination angle of Arase is larger than that of Van Allen Probes, Arase collected observations at higher L -shells up to L ∼ 10 . After March 2017, similar variations in plasma and waves were detected by Van Allen Probes and Arase. We describe plasma wave observations at longitudinally separated locations in space and geomagnetically-conjugate locations in space and on the ground. The results of instrument intercalibrations between the two missions are also presented. Arase continued its normal operation after the scientific operation of Van Allen Probes completed in October 2019. The combined Van Allen Probes (2012-2019) and Arase (2017-present) observations will cover a full solar cycle. This will be the first comprehensive long-term observation of the inner magnetosphere and radiation belts.

Direct measurements of two-way wave-particle energy transfer in a collisionless space plasma.

Particle acceleration by plasma waves and spontaneous wave generation are fundamental energy and momentum exchange processes in collisionless plasmas. Such wave-particle interactions occur ubiquitously in space. We present ultrafast measurements in Earth's magnetosphere by the Magnetospheric Multiscale spacecraft that enabled quantitative evaluation of energy transfer in interactions associated with electromagnetic ion cyclotron waves. The observed ion distributions are not symmetric around the magnetic field direction but are in phase with the plasma wave fields. The wave-ion phase relations demonstrate that a cyclotron resonance transferred energy from hot protons to waves, which in turn nonresonantly accelerated cold He+ to energies up to ~2 kilo-electron volts. These observations provide direct quantitative evidence for collisionless energy transfer in plasmas between distinct particle populations via wave-particle interactions.

Spatiotemporal treadmill gait measurements using a laser range scanner: feasibility study of the healthy young adults.

OBJECTIVE: Spatio-temporal parameters are typically used for gait analysis. Although these parameters are measured by sophisticated systems such as 3D motion capture system or optoelectronic bars, these systems cannot be deployed easily because of their high costs, large space requirements and elaborate set-up. The purpose of this study is to develope a system for measuring spatiotemporal gait parameters using a laser range scanner during treadmill gait. APPROACH: To calculate accurate spatiotemporal parameters, the differences between the laser range scanner measured values and the reference values obtained from a 3D motion capture system were investigated in thirty subjects. From measurements in time and position at foot contact/off, adjustments to compensate for the differences in time and position were derived. Then, to determine the validity of the proposed system, values from the proposed system and the reference system were compared in four additional subjects. MAIN RESULTS: The results indicate that the data from the laser range scanner demonstrate certain differences in time and position compared with reference values. However, when compensation values were introduced, each spatiotemporal parameter correlated well with the reference values. SIGNIFICANCE: This newer system is smaller, is easier to deploy and requires less training than the 3D motion capture system.

Application of X-ray microcomputed tomography in the characterization of irradiated nuclear fuel and material specimens.

X-ray microcomputed tomography (µCT) was applied in characterizing the internal structures of a number of irradiated materials, including carbon-carbon fibre composites, nuclear-grade graphite and tristructural isotropic-coated fuel particles. Local cracks in carbon-carbon fibre composites associated with their synthesis process were observed with µCT without any destructive sample preparation. Pore analysis of graphite samples was performed quantitatively, and qualitative analysis of pore distribution was accomplished. It was also shown that high-resolution µCT can be used to probe internal layer defects of tristructural isotropic-coated fuel particles to elucidate the resulting high release of radioisotopes. Layer defects of sizes ranging from 1 to 5 µm and up could be isolated by tomography. As an added advantage, µCT could also be used to identify regions with high densities of radioisotopes to determine the proper plane and orientation of particle mounting for further analytical characterization, such as materialographic sectioning followed by optical and electron microscopy. In fully ceramic matrix fuel forms, despite the highly absorbing matrix, characterization of tristructural isotropic-coated particles embedded in a silicon carbide matrix was accomplished using µCT and related advanced image analysis techniques.

Characterization of spontaneous malignant lymphomas in Japanese macaques (Macaca fuscata).

Lymphomas are common spontaneous tumors in nonhuman primates but remain poorly characterized in Japanese macaques (Macaca fuscata). This study examined 5 cases of spontaneous malignant lymphoma in Japanese macaques, focusing on the immunophenotypes and presence of simian lymphocryptoviruses, which are Epstein-Barr virus-related herpesviruses in nonhuman primates. The macaques with lymphoma were 5 to 28 years old, indicating that lymphomas develop over a wide age range. The common macroscopic findings were splenomegaly and enlargement of lymph nodes. Histologic and immunohistochemical analyses revealed that all cases were non-Hodgkin type and exhibited a T-cell phenotype, positive for CD3 but negative for CD20 and CD79α. The lymphomas exhibited diverse cellular morphologies and were subdivided into 3 types according to the World Health Organization classification. These included 3 cases of peripheral T-cell lymphoma, not otherwise specified; 1 case of T-cell prolymphocytic leukemia; and 1 case of an unclassifiable T-cell lymphoma. Positive signals were detected by in situ hybridization in 2 of the 4 examined cases using probes for the Epstein-Barr virus-encoded small RNA (EBER). Furthermore, the presence of M. fuscata lymphocryptovirus 2, a macaque homolog of Epstein-Barr virus, was demonstrated in EBER-positive cases by polymerase chain reaction amplification followed by direct sequencing. Immunohistochemistry using antibody to the Epstein-Barr virus-encoded nuclear antigen 2 was negative, even in the EBER-positive cases. The present study suggests that T-cell lymphoma is more common than B-cell lymphoma in Japanese macaques and that M. fuscata lymphocryptovirus 2 is present in some cases.

Downregulation of miRNA-31 induces taxane resistance in ovarian cancer cells through increase of receptor tyrosine kinase MET.

Ovarian cancer is one of the most aggressive female reproductive tract tumors. Paclitaxel (PTX) is widely used for the treatment of ovarian cancer. However, ovarian cancers often acquire chemotherapeutic resistance to this agent. We investigated the mechanism of chemoresistance by analysis of microRNAs using the ovarian cancer cell line KFr13 and its PTX-resistant derivative (KFr13Tx). We found that miR-31 was downregulated in KFr13Tx cells, and that re-introduction of miR31 re-sensitized them to PTX both in vitro and in vivo. miR-31 was found to bind to the 3'-UTR of mRNA of MET, and the decrease in MET correlated to higher sensitivity to PTX. Furthermore, co-treatment of KFr13Tx cells with MET inhibitors sensitized the tumor cells to PTX both in vitro and in vivo. In addition, lower levels of miR31 and higher expression of MET in human ovarian cancer specimens were significantly correlated with PTX chemoresistance and poor prognosis. This study demonstrated miR31-dependent regulation of MET for chemoresistance of ovarian cancer, raising the possibility that combination therapy with a MET inhibitor and PTX will increase PTX efficacy.

Hedgehog target genes: mechanisms of carcinogenesis induced by aberrant hedgehog signaling activation.

Hedgehog signaling is aberrantly activated in glioma, medulloblastoma, basal cell carcinoma, lung cancer, esophageal cancer, gastric cancer, pancreatic cancer, breast cancer, and other tumors. Hedgehog signals activate GLI family members via Smoothened. RTK signaling potentiates GLI activity through PI3K-AKT-mediated GSK3 inactivation or RAS-STIL1-mediated SUFU inactivation, while GPCR signaling to Gs represses GLI activity through adenylate cyclase-mediated PKA activation. GLI activators bind to GACCACCCA motif to regulate transcription of GLI1, PTCH1, PTCH2, HHIP1, MYCN, CCND1, CCND2, BCL2, CFLAR, FOXF1, FOXL1, PRDM1 (BLIMP1), JAG2, GREM1, and Follistatin. Hedgehog signals are fine-tuned based on positive feedback loop via GLI1 and negative feedback loop via PTCH1, PTCH2, and HHIP1. Excessive positive feedback or collapsed negative feedback of Hedgehog signaling due to epigenetic or genetic alterations leads to carcinogenesis. Hedgehog signals induce cellular proliferation through upregulation of N-Myc, Cyclin D/E, and FOXM1. Hedgehog signals directly upregulate JAG2, indirectly upregulate mesenchymal BMP4 via FOXF1 or FOXL1, and also upregulate WNT2B and WNT5A. Hedgehog signals induce stem cell markers BMI1, LGR5, CD44 and CD133 based on cross-talk with WNT and/or other signals. Hedgehog signals upregulate BCL2 and CFLAR to promote cellular survival, SNAI1 (Snail), SNAI2 (Slug), ZEB1, ZEB2 (SIP1), TWIST2, and FOXC2 to promote epithelial-to-mesenchymal transition, and PTHLH (PTHrP) to promote osteolytic bone metastasis. KAAD-cyclopamine, Mu-SSKYQ-cyclopamine, IPI-269609, SANT1, SANT2, CUR61414 and HhAntag are small-molecule inhibitors targeted to Smoothened, GANT58, GANT61 to GLI1 and GLI2, and Robot-nikinin to SHH. Hedgehog signaling inhibitors should be used in combination with RTK inhibitors, GPCR modulators, and/or irradiation for cancer therapy.

Disease-specific expression of VEGF and its receptors in AML cells: possible autocrine pathway of VEGF/type1 receptor of VEGF in t(15;17) AML and VEGF/type2 receptor of VEGF in t(8;21) AML.

Various angiogenic factors, such as vascular endothelial growth factor (VEGF) and an associated molecule, placenta growth factor (PlGF), are thought to be important for normal and malignant hematopoiesis. This study examined mRNA expression of VEGF, PlGF and receptors for these molecules in AML cells and identified the disease-specific patterns of expression. AML M3 having t(15;17) abnormality showed highest expression of VEGF and VEGF receptor type 1 (VEGFR1), suggesting the autocrine pathway of VEGF-VEGFR1. Then, t(8;21) AML demonstrated augmented expression of VEGF and VEGF receptor type 2 (VEGFR2), suggesting VEGF-VEGFR2 autocrine pathway. Then, addition of VEGFR2 kinase inhibitor in Kasumi-1, a t(8;21) AML cell line, resulted in marked inhibition of cell growth, although growth inhibitory effect of R2 kinase inhibitor to HL-60 was marginal. In addition, cell cycle analysis study showed S-phase cell population reduction by R2 kinase inhibitor in Kasumi-1, but not in HL-60. This observation is thought to be the rationale for novel molecular target therapy directed to angiogenic molecules.

Constrained genomic and conformational variability of the hypervariable region 1 of hepatitis C virus in chronically infected patients.

We analysed the genomic and conformational variability of the hypervariable region 1 (HVR1) of the hepatitis C virus (HCV) to evaluate the importance of its biological role. A total of 865 genotype 1b HVR1 subclones were collected from serially sampled sera in 11 patients with chronic hepatitis C, four of whom received interferon therapy. Consequently, 169 distinct sequences were examined for amino acid substitutions as well as hydrophilic or hydrophobic profile at each amino acid position within HVR1. Secondary structure of HVR1 was also predicted by the method of Robson in 90 distinct sequences from eight patients, including three interferon-treated patients. Some positions within the HVR1 were invariable or nearly so as to amino acid substitution. Hydrophilic or hydrophobic residues exclusively predominated at several positions. These constrained amino acid replacement and hydrophilic or hydrophobic profiles were conserved irrespective of interferon therapy, though the frequency of amino acid replacement was greater at almost all amino acid positions within the HVR1 in interferon-treated patients. The quasispecies of HCV showed various secondary structures of HVR1, but many sequences seemed to have common characteristics. beta sheet conformations around both the N-terminus and position 20 (numbered from the NH2 terminus of E2 envelope glycoprotein), and/or coil structures around the C-terminus of HVR1 could be identified. These results suggest that HVR1 amino acid replacements are strongly constrained by a well-ordered structure, in spite of being tolerant to amino acid substitutions, and imply an important biological role of the HVR1 protein in HCV replication.

Expression of endothelial cell-associated molecules in AML cells.

Recently, it has been clarified that interaction between hematopoietic cells and endothelial cells is important in normal hematopoiesis and leukemogenesis. In this study, we examined the relationship between AML cells and endothelial cells by analyzing the expression profile of angiogenic factors, angiopoietin-1 (Ang-1), Ang-2, Tie-2 (a receptor for angiopoietins) and vascular endothelial growth factor (VEGF). Our results demonstrated that CD7(+)AML expressed Ang-2 mRNA frequently and integrin-family adhesion molecules (CD11c and CD18) intensively, suggesting the close correlation with endothelial cells. On the other hand, in t(8;21) AML cells, expression of Ang-2 was infrequent and expression of integrin-family adhesion molecules (CD11b, CD11c and CD18) was weak, suggesting the sparse association with endothelial cells. As for CD7(+)AML cells, despite the frequent and intense expression of endothelial cell-associated molecules (such as Ang-2, CD11c and CD18), intensity of Tie-2 expression was quite low (P < 0.05). Ang-2 expressed in CD7(+)AML cells is not considered to act in an autocrine fashion, but to work on endothelial cells to "feed" leukemic cells. Although Ang-2 is recognized as a natural antagonist for Tie-2, our data presented here suggested the alternative role of Ang-2 in the relationship between endothelial cells and leukemia cells, at least in a subset of leukemia such as CD7(+)AML. These results were supported by the study using AML cell lines, KG-1 (CD7 negative) and its subline KG-1a (CD7 positive); KG-1 had mRNA expression profile of Ang-1(+)Ang-2(-)Tie-2(+), while KG-1a showed Ang-1(+)Ang-2(+)Tie-2(-). These difference in the expression profile of angiogenic factors between CD7(+)AML and t(8;21)AML may explain the characteristic morphological features of these leukemias (CD7(+)AML as blastic type and t(8;21)AML as differentiative type).

Arterial ketone body ratio for the assessment of the severity of illness in pediatric patients following cardiac surgery.

PURPOSE: The purpose of this study was to assess whether the arterial ketone body ratio (AKBR) can be effectively used to evaluate the severity of illness in children following cardiac surgery. MATERIALS AND METHODS: AKBR was measured in 157 consecutive pediatric patients following heart surgery on the odd numbers of postoperative days. The relationship between AKBR and patient outcome was analyzed using the data of 141 patients with cardiopulmonary bypass. RESULTS: Initial AKBR was frequently lower than 1.0, and this was associated with the increases in total ketone body counts. Insufficient glucose metabolism appeared to contribute to the low initial AKBR. As a result, the specificity of initial AKBR as a mortality predictor was lower than that of initial blood lactate. In the sequential analysis of AKBR for the 48 patients with PICU stay longer than 5 days, patients showing a sustained lower level <1.0 had significantly higher development of organ dysfunction (liver, heart) and greater mortality (56%). CONCLUSIONS: Sustained postoperative decrease in AKBR <1.0 represents lethal outcome. The analysis of AKBR trend in combination with a measurement of blood lactate level in early postoperative period appears to be useful for the assessment of the severity of illness in pediatric patients following heart surgery.

[The number and form of cerebellar fastigial neurons projecting to the thalamic suprageniculate nucleus in cats studied using WGA-HRP tracing method].

This study was designed to investigate the number and form of cerebellar fastigial neurons projecting to the suprageniculate nucleus(Sg) by using retrograde axonal transport of wheat germ agglutinin-horseradish peroxidase(WGA-HRP). Six adult cats(weighing 2.5-3.5 kg) were anesthetized with ketamine hydrochloride(30 mg/kg, i.m.) and sodium pentobarbital(15 mg/kg, i.p.). In experiments using injected WGA-HRP in the Sg, retrogradely labeled neurons by WGA-HRP were found only in the caudal part of the bilateral fastigial nucleus (Ft) with ipsilateral predominance, and the ratio of labeled neurons in the contralateral Ft to that in the ipsilateral Ft in all cats was 496:670(1:1.35). Five types of Ft-Sg neurons were distinguished morphologicaly. Of the 246 labeled neurons that could be characterized, 6.1% were large stellate neurons, 43.5% medium stellate neurons, 24.0% bipolar neurons, 20.3% triangular neurons, and 6.1% granular neurons. Thus, we concluded that a mixed population of Ft neurons projects to the Sg. In summary, as shown in the Figure 4 the Ft-Sg connection is an important pathway joining two closed circuits, and can be part of the extrageniculate visual system. We also speculate that the Ft-Sg connections may have a role in sending visual modulating information.

Two domains of Nrf2 cooperatively bind CBP, a CREB binding protein, and synergistically activate transcription.

BACKGROUND: Nrf2 belongs to the Cap-N-Collar (CNC) transcription factor family and is essential for the antioxidant responsive element (ARE)-mediated expression of a group of detoxifying and antioxidant genes. The forced expression of Nrf2 in mammalian cells activates the expression of target genes through the ARE, with Nrf2 showing the highest transactivation activity among the CNC family of transcription factors. To elucidate the molecular mechanisms generating this potent transactivation activity, we examined the functions of the domains within Nrf2. RESULT: We found that Nrf2 contains two transcription activation domains, Neh4 and Neh5, which act synergistically to attain maximum a activation of reporter gene expression. Neh4 and Neh5 both individually and cooperatively bind to CBP (CREB (cAMP Responsive Element Binding protein) Binding Protein). In fact, the specific inhibitor of CBP, adenovirus E1A protein, significantly reduced Nrf2 activity. Importantly, the CBP-binding activity of Nrf2 deletion mutants positively correlated with their transactivation activity. Neh5 contains a motif which is commonly conserved among the CNC factors, whereas Neh4 contains the novel CBP-interacting motif recently identified in p53 and E2F. CONCLUSIONS: Our results indicate that Nrf2 exploits the cooperative binding of two independent transactivation domains to CBP in the acquisition of a potent transactivation activity.

Intraosseous ganglion in the first metacarpal bone.

Intraosseous ganglia occur most frequently in the long bones of the lower limbs, particularly in the medial malleolus of the tibia. They usually appear as radiographically well circumscribed juxta-articular cystic lesions, containing myxoid fibrous tissue histologically. Intraosseous ganglia in the hand are very rare. Most reported cases have involved the carpal bones, in particular the lunate and scaphoid. To our knowledge, the present case is the third report of an intraosseous ganglion appearing in the first metacarpal bone; it arose in a patient who had been on dialysis for 25 years, mimicking amyloidosis of bone.

A possible role of hypervariable region 1 quasispecies in escape of hepatitis C virus particles from neutralization.

We examined serial changes in the hypervariable region 1(HVR1) quasispecies both in immune and nonimmune complexed hepatitis C virus (HCV) particles from 12 patients with chronic hepatitis C to elucidate the mechanism by which genetic diversification of HCV during the course of infection allows escape of virus from the humoural immune response. Immune and nonimmune complexes were separated by differential flotation centrifugation and immunoprecipitation, and their HVR1 quasispecies were determined by subcloning and sequencing. The presence of a specific antibody against a specific viral clone in serum was examined in two patients by Western blotting of the corresponding recombinant HVR1 protein. The distribution of HVR1 quasispecies in both immune and nonimmune complexes conspicuously changed over time in most of the patients studied. In seven patients, various HCV clones serially shifted from nonimmune complexes to immune complexes. In four of them, a group of clones with similar HVR1 sequences to each other remained predominant in nonimmune complexes, whereas minor clones with sequences considerably divergent from the predominant clones shifted from nonimmune complexes to immune complexes. These results suggest a mechanism for persistent infection of HCV, in which major HCV clones escape from neutralization by anti-HVR1 antibodies by generating considerably divergent minor 'decoy' clones which may be preferentially neutralized.

Salt-inducible kinase is involved in the ACTH/cAMP-dependent protein kinase signaling in Y1 mouse adrenocortical tumor cells.

The involvement of salt-inducible kinase, a recently cloned protein serine/threonine kinase, in adrenal steroidogenesis was investigated. When Y1 mouse adrenocortical tumor cells were stimulated by ACTH, the cellular content of salt-inducible kinase mRNA, protein, and enzyme activity changed rapidly. Its level reached the highest point in 1-2 h and returned to the initial level after 8 h. The mRNA levels of cholesterol side-chain cleavage cytochrome P450 and steroidogenic acute regulatory protein, on the other hand, began to rise after a few hours, reaching the highest levels after 8 h. The salt-inducible kinase mRNA level in ACTH-, forskolin-, or 8-bromo-cAMP-treated Kin-7 cells, mutant Y1 with less cAMP-dependent PKA activity, remained low. However, Kin-7 cells, when transfected with a PKA expression vector, expressed salt-inducible kinase mRNA. Y1 cells that overexpressed salt-inducible kinase were isolated, and the mRNA levels of steroidogenic genes in these cells were compared with those in the parent Y1. The level of cholesterol side-chain cleavage cytochrome P450 mRNA in the salt-inducible kinase-overexpressing cells was markedly low compared with that in the parent, while the levels of Ad4BP/steroidogenic factor-1-, ACTH receptor-, and steroidogenic acute regulatory protein-mRNAs in the former were similar to those in the latter. The ACTH-dependent expression of cholesterol side-chain cleavage cytochrome P450- and steroidogenic acute regulatory protein-mRNAs in the salt-inducible kinase-overexpressing cells was significantly repressed. The promoter activity of the cholesterol side-chain cleavage cytochrome P450 gene was assayed by using Y1 cells transfected with a human cholesterol side-chain cleavage cytochrome P450 promoter-linked reporter gene. Addition of forskolin to the culture medium enhanced the cholesterol side-chain cleavage cytochrome P450 promoter activity, but the forskolin-dependently activated promoter activity was inhibited when the cells were transfected with a salt-inducible kinase expression vector. This inhibition did not occur when the cells were transfected with a salt-inducible kinase (K56M) vector that encoded an inactive kinase. The salt-inducible kinase's inhibitory effect was also observed when nonsteroidogenic, nonAd4BP/steroidogenic factor-1 -expressing, NIH3T3 cells were used for the promoter assays. These results suggested that salt-inducible kinase might play an important role(s) in the cAMP-dependent, but Ad4BP/steroidogenic factor-1-independent, gene expression of cholesterol side-chain cleavage cytochrome P450 in adrenocortical cells.

The effect of introduction of universal childhood hepatitis B immunization in South Africa on the prevalence of serologically negative hepatitis B virus infection and the selection of immune escape variants.

The effect of universal hepatitis B vaccination on the prevalence of serologically negative hepatitis B virus infection (HBV) and the emergence of HBsAg variants is unknown. We prospectively studied two different cohorts of 12--24 month old children in South Africa. They consisted of the unvaccinated children (n=459) born before the introduction of universal vaccination and the vaccinated children (n=1213) between 1 and 2 years after the introduction of the vaccination program. The frequency of detecting HBV DNA by PCR was reduced from 6.5% in unvaccinated children to 0.3% in vaccinated children (P<0.00001). There were no unique amino acid substitutions within the major hydrophilic region of the S sequence in both pre- and post-vaccination samples. Universal childhood vaccination reduced the frequency of serologically negative HBV infection and did not necessarily lead to selection of escape variants.

Histamine production by cultured microglial cells of the mouse.

We previously reported that cells other than mast cells or neurons could synthesize histamine in response to lipopolysaccharide (LPS) or interleukin 1beta in the rat brain. To identify the responsible cells, we examined histidine decarboxylase (HDC) activity and the expression of HDC mRNA in GMI 6-3 mouse microglial cells. Both the activity and mRNA for HDC in GMI 6-3 cells were induced by LPS treatment, and the induction was sensitive to calmodulin-dependent kinase II inhibitor, KN62. These findings indicate that microglia is a third cell type producing histamine in the brain.

Golgi-localizing, gamma-adaptin ear homology domain, ADP-ribosylation factor-binding (GGA) proteins interact with acidic dileucine sequences within the cytoplasmic domains of sorting receptors through their Vps27p/Hrs/STAM (VHS) domains.

GGA (Golgi-localizing, gamma-adaptin ear homology domain, ARF-binding) proteins are potential effectors of ADP-ribosylation factors, are associated with the trans-Golgi network (TGN), and are involved in protein transport from this compartment. By yeast two-hybrid screening and subsequent two-hybrid and pull-down analyses, we have shown that GGA proteins, through their VHS (Vps27p/Hrs/STAM) domains, interact with acidic dileucine sequences found in the cytoplasmic domains of TGN-localized sorting receptors such as sortilin and mannose 6-phosphate receptor. A mutational analysis has revealed that a leucine pair and a cluster of acidic residues adjacent to the pair are mainly responsible for the interaction. A chimeric receptor with the sortilin cytoplasmic domain localizes to the TGN, whereas the chimeric receptor with a mutation at the leucine pair or the acidic cluster is mislocalized to punctate structures reminiscent of early endosomes. These results indicate that GGA proteins regulate the localization to or exit from the TGN of the sorting receptors.