Childhood anhedonia symptoms and stressful life events predict the development of reward-related brain activity across adolescence.

The reward positivity (RewP) is an event-related potential that indexes reinforcement learning and reward system activation. The RewP has been shown to increase across adolescence; however, most studies have examined the RewP across two assessments, and no studies have examined within-person changes across adolescence into young adulthood. Moreover, the RewP has been identified as a neurobiological risk factor for adolescent-onset depression, but it is unclear whether childhood psychosocial risk factors might predict RewP development across adolescence. In a sample of 317 8- to 14-year-old girls (Mage = 12.4, SD = 1.8), the present study examined self-report measures of depression symptoms and stressful life events at baseline and the ΔRewP during the doors guessing task across three timepoints. Growth modeling indicated that, across all participants, the ΔRewP did not demonstrate linear change across adolescence. However, baseline anhedonia symptoms predicted within-person changes in the ΔRewP, such that individuals with low anhedonia symptoms demonstrated a linear increase in the ΔRewP, but individuals with high anhedonia symptoms had no change in the ΔRewP across adolescence. Similar patterns were observed for stressful life events. The present study suggests that childhood risk factors impact the development of reward-related brain activity, which might subsequently increase risk for psychopathology.

A postpartum haemorrhage package with condom uterine balloon tamponade: a prospective multi-centre case series in Kenya, Sierra Leone, Senegal, and Nepal.

OBJECTIVE: To evaluate the effectiveness and safety of an ultra-low-cost uterine balloon tamponade package (ESM-UBT™) for facility-based management of uncontrolled postpartum haemorrhage (PPH) in Kenya, Sierra Leone, Senegal, and Nepal. DESIGN: Prospective multi-centre case series. SETTING: Facilities in resource-scarce areas of Kenya, Sierra Leone, Nepal, and Senegal. POPULATION: Women with uncontrolled postpartum haemorrhage in 307 facilities across the four countries. METHODS: A standardised ESM-UBT package was implemented in 307 facilities over 29 months (1 September 2012 to 1 February 2015). Data were collected via a multi-pronged approach including data card completion, chart reviews, and provider interviews. Beginning in August 2014, women who had previously undergone UBT placement were sought and queried regarding potential complications associated with UBT use. MAIN OUTCOME MEASURES: All-cause survival, survival from PPH, and post-UBT use complications (surgery, hospitalisation, antibiotics for pelvic infection) associated with UBT use. RESULTS: 201 UBTs were placed for uncontrolled vaginal haemorrhage refractory to all other interventions. In all, 38% (71/188) of women were either unconscious or confused at the time of UBT insertion. All-cause survival was 95% (190/201). However, 98% (160/163) of women survived uncontrolled PPH if delivery occurred at an ESM-UBT online facility. One (1/151) potential UBT-associated complication (postpartum endometritis) was identified and two improvised UBTs were placed in women with a ruptured uterus. CONCLUSIONS: These pilot data suggest that the ESM-UBT package is a clinically promising and safe method to arrest uncontrolled postpartum haemorrhage and save women's lives. The UBT was successfully placed by all levels of facility-based providers. Future studies are needed to further evaluate the effectiveness of ESM-UBT in low-resource settings. TWEETABLE ABSTRACT: Evidence for ESM-UBT as a clinically promising and safe method to arrest uncontrolled PPH and save women's lives.

Youth in crisis in the Middle East and North Africa: a systematic literature review and focused landscape analysis.

Recent political and demographic factors have exposed the vulnerability of the youth in the Middle East and North Africa (MENA) region. This study aimed to elucidate the current needs, activities, stakeholders and solutions related to at-risk youth and young adults in the MENA region. A systematic literature review was conducted of the peer-reviewed and grey literature. This was complemented by an in-region landscape analysis involving key-informant interviews and focus group discussions. After extensive screening of 1160 unique articles, 275 articles were considered relevant to this study. Of these 275, 145 (52.7%) were related to health (64.8% of these related to mental health), 101 (36.7%) to livelihood, 87 (31.6%) to violence prevention and 68 (24.7%) to education. Important themes and challenges identified in the literature and discussions included the MENA region's growing youth bulge; youth unemployment; critical gender gaps; and the impact of conflict on livelihoods, education and health, especially mental health.

Familial risk for distress and fear disorders and emotional reactivity in adolescence: an event-related potential investigation.

BACKGROUND: The late positive potential (LPP) is an event-related potential component that is sensitive to the motivational salience of stimuli. Children with a parental history of depression, an indicator of risk, have been found to exhibit an attenuated LPP to emotional stimuli. Research on depressive and anxiety disorders has organized these conditions into two empirical classes: distress and fear disorders. The present study examined whether parental history of distress and fear disorders was associated with the LPP to emotional stimuli in a large sample of adolescent girls. METHOD: The sample of 550 girls (ages 13.5-15.5 years) with no lifetime history of depression completed an emotional picture-viewing task and the LPP was measured in response to neutral, pleasant and unpleasant pictures. Parental lifetime history of psychopathology was determined via a semi-structured diagnostic interview with a biological parent, and confirmatory factor analysis was used to model distress and fear dimensions. RESULTS: Parental distress risk was associated with an attenuated LPP to all stimuli. In contrast, parental fear risk was associated with an enhanced LPP to unpleasant pictures but was unrelated to the LPP to neutral and pleasant pictures. Furthermore, these results were independent of the adolescent girls' current depression and anxiety symptoms and pubertal status. CONCLUSIONS: The present study demonstrates that familial risk for distress and fear disorders may have unique profiles in terms of electrocortical measures of emotional information processing. This study is also one of the first to investigate emotional/motivational processes underlying the distress and fear disorder dimensions.

Perinatal health care in a conflict-affected setting: evaluation of health-care services and newborn outcomes at a regional medical centre in Iraq.

A field-based assessment was conducted to assess maternal and newborn health-care services, perinatal and newborn outcomes and associated risk factors at Bint Al-Huda Maternal and Newborn Teaching Hospital, a large referral hospital in southern Iraq. The multi-method approach used interviews, discussions, observation and review of perinatal and newborn outcome data. There is limited assessment of maternal vital signs, labour pattern, fetal response, and complications during pregnancy and labour. Perinatal and neonatal mortality rates are 27.4/1000 births and 30.9/1000 live births respectively. Associated neonatal mortality factors were gestational age < 37 weeks, male sex, birth weight < 2.5 kg, maternal age > 35 years, rural maternal residence and vaginal delivery. Improving birth outcomes in southern Iraq requires evidence-based clinical guidelines, additional supplies and equipment, quality improvement initiatives and in-service training.

Development of a community-based maternal, newborn and child emergency training package in South Sudan.

OBJECTIVE: To develop an evidence-based maternal, newborn and child emergency training package for community-based frontline health workers (FHWs) in post-conflict South Sudan. METHODS: In partnership with the new Republic of South Sudan, a multimodal needs assessment was conducted through purposive sampling, involving key informant interviews, focus group discussions, provider knowledge assessments and facility surveys. Data were analyzed using traditional qualitative techniques and compared with existing training programmes and curricula. These findings informed the development and implementation of the novel training approach. RESULTS: The needs assessment involved 33 FHWs, eight diverse health facilities in Eastern Equatoria, and stakeholders within 18 governmental and non-governmental organizations. Significant consensus emerged regarding the need for greater capacity among previously untrained FHWs. A maternal, newborn and child health training package was developed that included: (1) a participatory training course taught through a 'training of trainers' approach; (2) nine different pictorial action-based checklists covering basic management and referral of maternal, newborn and child emergencies; and (3) essential setting-appropriate equipment. CONCLUSION: A novel maternal, newborn and child survival package was developed for previously untrained and illiterate FHWs in South Sudan. It is hoped that this approach will build community-based capacity in resource-limited settings while greater capacity is being developed for facility-based deliveries by skilled birth attendants.

First Report of Alfalfa mosaic virus and Soybean dwarf virus on Soybean in North Dakota.

Soybean (Glycine max L.) is the major oilseed crop in North Dakota, with production concentrated in the eastern half of the state. Only one virus, Soybean mosaic virus, has been reported from soybean in North Dakota (4). In July and August of 2010, 200 soybean fields from 25 counties were surveyed for Alfalfa mosaic virus (AMV) and Soybean dwarf virus (SbDV). AMV and SbDV have been detected infecting soybean in multiple Midwestern states and are reported to reduce yields in soybean (1,3). Each field was sampled with a grid pattern across the area with at least 8 km between fields. From each field, leaves were collected from 20 plants without regard for symptoms along a transect of approximately 170 m. Leaves from each field were bulked and sap was extracted in phosphate buffer and stored at -80°C until tested using double-antibody sandwich (DAS)-ELISA with positive controls and reagents and protocols from Agdia Inc. (Elkhart, IN). Using DAS-ELISA, AMV was detected in eight of the 200 soybean fields. For sequence-based virus detection, total RNA was extracted from all field samples using a Qiagen RNeasy Plant Mini Kit (Germantown, MD), pooled, depleted of ribosomal RNA (RiboZero Epicentre, Madison, WI), reverse transcribed, sequenced using an Illumina HiSeq2000 (San Diego, CA), and compared to all available viral amino acid and nucleotide sequences. The analysis detected AMV and SbDV sequences in the pool of 200 fields. The presence of AMV and SbDV was confirmed by quantitative real-time reverse transcription (qRT)-PCR (1,3). For AMV, total RNA extracted from bulked leaves from each of the 200 fields was tested using AMVspecific primers (5'-ATGCTACCCAGGCATGTATATTT-3' and 5'-GCTGCATCTTTCGCCAGAA-3') and a FAM-labeled minor-groove binding TaqMan probe (5'-TGGACGTTACCCCCGGA-3'). One field sample from Cass county positive for AMV by ELISA was also positive for AMV by qRT-PCR, confirming the presence of AMV in the field sample. For SbDV, an RNA pool representing all 200 fields, subpools, and individual field samples was analyzed by qRT-PCR (1) and DAS-ELISA. One field sample from Grand Forks County tested positive for SbDV by qRT-PCR and DAS-ELISA, confirming the presence of SbDV in the field sample. Because leaf samples were collected and pooled prior to analysis, the symptom phenotypes of individual field plants could not be correlated with positive ELISA or qRT-PCR results. AMV was reported by the American Phytopathological Society Virus Working Group (2007 to 2008) to be widely prevalent in North Dakota, but we found no peer-reviewed reports of verified AMV identification on any crop in the state. To our knowledge, this is the first confirmed report of AMV and SbDV infecting soybean in North Dakota. Serious infestations by the soybean aphid, Aphis glycines, requiring chemical control, have occurred in recent years in North Dakota. Because A. glycines is a vector for both viruses (1,2), the distribution, incidence, and agronomic impact of AMV and SbDV could be affected in years when A. glycines infestations are high. In addition, AMV is seedborne in soybean and may cause seed mottling, a concern for the food-grade soybean industry where production is primarily for export. References: (1) V. D. Damsteegt et al. Plant Dis. 95:945, 2011 (2) J. H. Hill et al. Plant Dis. 85:561, 2001. (3) H. A. Hobbs et al. Plant Health Progress doi:10.1094/PHP-2010-0827-01-BR, 2010. (4) B. D. Nelson and L. L. Domier. Plant Dis. 93:760, 2009.

First Confirmed Report of Sugar Beet Cyst Nematode, Heterodera schachtii, in North Dakota.

Sugar beet (Beta vulgaris L.) and canola (Brassica napus L.) are major crops in North Dakota, with sugar beet production primarily in the eastern part of the state in the Red River Valley and canola production across the northern half of the state. Both crops are hosts of sugar beet cyst nematode (SBCN), Heterodera schachtii Schmidt. In April 2011, soil samples were collected from four sugar beet fields belonging to three growers who believed the fields were infested with SBCN. The fields were located in a 65-km2 area in the Yellowstone Valley of western North Dakota. Cysts were extracted by sieving and Heterodera-like cysts with eggs were observed in all four soil samples. Population densities in the four fields ranged from 100 to 1,750 eggs/100 cm3 soil. Sugar beet seedlings (cv. M832224) were grown in a potting mix for 6 weeks in the greenhouse and then transferred to conetainers (type D40; volume 656 ml) containing autoclaved river sand. Conetainers were placed in sand in plastic pots immersed in a water bath at 27°C. Three plants were each infested with 800 eggs from field No. 2. After 55 days of incubation, the average number of females was 115 per plant. A similar experiment was conducted with canola cvs. Hyclass 940, Caliber 30, and Westar, which were inoculated with 500 eggs each from field No. 2. After 53 days of incubation, there was an average of 39, 20, and 30 females for each respective cultivar. Flask-shaped cysts (n = 26) from canola roots were light to dark brown; the vulval cone was ambifinestrate with dark brown, molar-shaped bullae positioned underneath the vulval bridge. Body length (excluding neck) ranged from 600 to 850 µm (mean 701.2 µm); body width, 350 to 580 µm (mean 469.2 µm); and length/width ratio, 1.2 to 1.8 (mean 1.5). Second-stage juvenile (J2) (n = 21) body length ranged from 400 to 485 µm (mean 437.1 µm); stylet length was 25 µm (no variation) with forwardly directed knobs; conical tail with rounded tip ranged from 37.5 to 55.0 µm long (mean 46.6 µm) with hyaline region from 20.0 to 32.5 µm (mean 27.3 µm); and lateral field presented four incisures. These morphometrics were used to identify H. schachtii according to Subbotin et al. (4). Confirmation of identification was by amplification and sequencing of a 28S rDNA gene fragment (1) from individual females (GenBank Accession No. JQ040526), which was 100% identical to H. schachtii 28S rDNA sequence (GenBank Accession No. GU475088). To our knowledge, this is the first confirmed report of H. schachtii in North Dakota. A 1958 report of SBCN in North Dakota (2) was not subsequently confirmed (3). Because there is extensive canola production across the northern part of the state bordering western and eastern sugar beet- production areas, canola may serve as a bridge for movement of SBCN from west to east. SBCN is a potential threat to these two important crops. References: (1) A. Amiri et al. Eur. J. Plant Pathol. 108:497, 2002. (2) F. Caveness. J. Sugar Beet Res. 10:544, 1958. (3) P. Donald and R. Hosford. Plant Dis. 64:45, 1980. (4) S. A. Subbotin et al. Systematics of Cyst Nematodes (Nematoda: Heteroderinae). Nematology Monographs and Perspectives. Vol. 8B. Brill, The Netherlands. 2010.

Characterization of the hepatic and splenic immune status and immunoglobulin synthesis in aged male mice lacking the peroxisome proliferator-activated receptor-alpha (PPARα).

It is now well established that the nuclear receptor peroxisome proliferator-activated receptor-alpha (PPARα) is expressed in different types of immune cells and plays a pivotal role in the regulation of age-related production of inflammatory cytokines. However, the role(s) of this receptor in the regulation of immune cell homoeostasis in ageing non-lymphoid and lymphoid organs has not yet been resolved. We examine this issue here by evaluating the hepatic and splenic immune status and immunoglobulin (Ig) production in male PPARα-null mice and their wild-type littermates at one and 2 years of age. In comparison with the age-matched control animals, PPARα-null mice exhibited age-related elevations in the numbers of total, as well as of phenotypically distinct subpopulations of intrahepatic immune cells (IHIC) and splenocytes. Moreover, at 2 years of age, these alterations in hepatic immune cells were accompanied by significant increases in hepatic levels of the pro-inflammatory cytokines tumour necrosis factor-alpha (TNF-α), interleukin-6 (IL-6) and interferon-gamma (IFN-γ), in combination with the development of hepatic inflammatory loci containing mixtures of leucocytes. Alterations in splenocytes of old PPARα-null mice were also accompanied by increases in cellularity of both white and red pulps of the spleen. Furthermore, these same animals exhibited pronounced increases in the numbers of splenic plasma cells and enhanced production of Ig of different isotypes, including IgG1, IgG2a and IgE. Thus, our findings indicate that upon ageing, PPARα plays a crucial role in regulating the total numbers, compositions and functions of immune cells in both lymphoid and non-lymphoid immune organs of mice.

Isolation of murine intrahepatic immune cells employing a modified procedure for mechanical disruption and functional characterization of the B, T and natural killer T cells obtained.

Intrahepatic immune cells (IHIC) are known to play central roles in immunological responses mediated by the liver, and isolation and phenotypic characterization of these cells is therefore of considerable importance. In the present investigation, we developed a simple procedure for the mechanical disruption of mouse liver that allows efficient isolation and phenotypic characterization of IHIC. These cells are compared with the corresponding cells purified from the liver after enzymatic digestion with different concentrations of collagenase and DNase. The mechanical disruption yielded viable IHIC in considerably greater numbers than those obtained following enzymatic digestion. The IHIC isolated employing the mechanical disruption were heterogeneous in composition, consisting of both innate and adaptive immune cells, of which B, T, natural killer (NK), NK T cells, granulocytes and macrophages were the major populations (constituting 37.5%, 16.5%, 12.1%, 7.9%, 7.9% and 7.5% of the total number of cells recovered respectively). The IHIC obtained following enzymatic digestion contained markedly lower numbers of NK T cells (1.8%). The B, T and NK T cells among IHIC isolated employing mechanical disruption were found to be immunocompetent, i.e. they proliferated in vitro in response to their specific stimuli (lipopolysaccharide, concanavalin A and alpha-galactosylceramide respectively) and produced immunoglobulin M and interferon-gamma. Thus, the simple procedure for the mechanical disruption of mouse liver described here results in more efficient isolation of functionally competent IHIC for various types of investigation.

First Report of Soybean mosaic virus on Soybean in North Dakota.

Soybean, Glycine max L, is grown on 1,420,000 ha in North Dakota and is the most important oilseed crop in the state. Viruses in soybean have not previously been reported from North Dakota (2). In July and August of 2007, 64 soybean fields in Cass, Richland, and Sargent counties in southeastern North Dakota were surveyed for Soybean mosaic virus (SMV). These counties have a high concentration of soybean hectares, a long history of soybean production, and soybean aphid infestations that were observed in 2004 and 2006. Fields were sampled with a grid pattern across the area with at least 8 km (5 miles) between fields. A transect of approximately 60 m through each field was made and 20 leaves were collected at random. Sap was extracted in phosphate buffer and stored at -80°C until tested first using double antibody sandwich (DAS)-ELISA with positive controls and reagents and protocol from Agdia Inc. (Elkhart, IN). Using DAS-ELISA, SMV was detected in 19 of the 64 soybean fields sampled. To confirm the presence of SMV, 12 samples that were positive for SMV by DAS-ELISA also were tested by reverse transcription (RT)-PCR. RNA was extracted from sap by a Qiagen RNeasy Plant Mini Kit (Germantown, MD), reverse transcribed, and amplified with SuperScrip III Platinum SYBR Green One-Step qRT-PCR Kit (Invitrogen Inc., Carlsbad, CA) and SMV-specific primers (5'-TTCAGCACAATGGGTGAGGATG-3' and 5'-AATTCTGTGTGGCTTGATGTTGC-3') (1). Eight of the twelve ELISA-positive samples were positive for SMV by RT-PCR, confirming the presence of SMV in the samples. To our knowledge, this is the first report of SMV infecting soybean in North Dakota. References: (1) L. L. Domier et al. (Abstr.). Phytopathology 98(suppl.):S47, 2008. (2) B. D. Nelson and G. Danielson. (Abstr.). Phytopathology 95(suppl.):S164, 2005.

Pathotypes, Distribution, and Metalaxyl Sensitivity of Phytophthora sojae from North Dakota.

Phytophthora root rot, caused by Phytophthora sojae, is the most important disease of soybean (Glycine max) in North Dakota. Because of the expansion of soybean hectares and appearance of disease on cultivars with resistance genes, we investigated the pathotypes, distribution, and metalaxyl sensitivity of P. sojae in North Dakota. Soil from 347 soybean fields in 20 counties in eastern North Dakota was collected between 2002 and 2004, and P. sojae was baited from the soil with the susceptible cultivar McCall. The virulence phenotype of each isolate was determined on eight differentials, and all isolates were tested for sensitivity to metalaxyl incorporated into V8 agar. The pathogen was recovered from 80 fields located in five counties. Sixteen pathotypes, which included 14 known races and two previously reported pathotypes that had not been assigned a race, were identified out of 157 isolates. A single pathotype was recovered from 61 fields, 2 pathotypes from 14 fields, 3 pathotypes from 4 fields, and 4 pathotypes from 1 field. Pathotypes with virulence phenotypes 1a,1c,7 (race 4; 39%) and 1a,7 (race 3; 28%) were the most common, representing 67% of the total isolates. One or both of these pathotypes was found in 79% of the fields where P. sojae was recovered. Seven of the 157 isolates showed limited growth on metalaxyl after 14 days of incubation. In the past 10 years, the number of pathotypes of P. sojae in North Dakota has increased from 4 to 16, and pathotypes have developed that can attack the three most common resistance genes found in soybean cultivars for the region.

Thyroid hormone regulation of mitochondrial function. Comments on the mechanism of signal transduction.

Thyroid hormone exerts two types of effect on mitochondria. The first of these is a rapid activation of respiration which takes place within minutes after hormone injection, and is preserved in isolated mitochondria. The second response occurs after 1 to several days of injection and leads to mitochondrial biogenesis and increases in mitochondria mass. The hormone signal for these two responses involves either triiodothyronine (T3)-responsive nuclear genes or a direct action of T3 at mitochondria binding sites.

Hexokinase bound to rat brain mitochondria uses externally added ATP more efficiently than internally generated ATP.

The rates of glucose phosphorylation by bound hexokinase were investigated in mitochondria isolated from rat brain. Initial rates obtained either with ATP generated from oxidative phosphorylation or with ATP added externally were compared. Our results show that the external ATP supports a 2-3-fold higher hexokinase activity than does ATP generated by oxidative phosphorylation under Stage 3 conditions. ATP formed by mitochondrial creatine kinase in the presence of creatine phosphate also supports higher initial rates of glucose phosphorylation than does oxidative phosphorylation. The data suggest that concentrations of ATP present in the cytosol of normal tissue will probably maintain higher rates of glucose phosphorylation than ATP being exported directly from the mitochondrial matrix at maximal State 3 rates.

Alkali-induced reduction of the beta-cytochromes in purified complex III from beef heart mitochondria.

Approx. 40-50% of the cytochrome b in purified Complex III is reduced by ascorbate plus N,N,N',N'-tetramethyl-p-phenylenediamine or phenazine methosulfate at neutral pH. The remaining cytochrome b, including cytochrome b-565, is reduced by increasing the pH. The apparent pK for this reduction is between pH 10 and 11, and is more than two pH units higher than a similar alkali-induced transition in Mg-ATP particles. Alkali-induced reduction of cytochrome b occurs concomitantly with the exposure of hydrophobic tyrosine and tryptophan residues to a more hydrophilic environment. The relationship of these findings to the presence of a substrate accessibility barrier in Complex III is discussed.

Adenylate kinase is a source of ATP for tumor mitochondrial hexokinase.

It has been proposed that hexokinase bound to mitochondria occupies a preferred site to which ATP from oxidative phosphorylation is channeled directly (Bessman, S. (1966) Am. J. Medicine 40, 740-749). We have investigated this problem in isolated Zajdela hepatoma mitochondria. Addition of ADP to well-coupled mitochondria in the presence of an oxidizable substrate initiates the synthesis of glucose 6-phosphate via bound hexokinase. This reaction is only partially inhibited by oligomycin, carboxyatractyloside, carbonyl cyanide m-chlorophenylhydrazone (CCCP) or any combination of these, suggesting a source of ATP in addition to oxidative phosPhorylation. This source appears to be adenylate kinase, since Ado2P5, an inhibitor of the enzyme, suppresses hexokinase activity by about 50% when added alone or suppresses activity completely when added together with any of the inhibitors of oxidative phosphorylation. Ado2P5 does not uncouple oxidative phosphorylation nor does it inhibit ADP transport (state 3 respiration) or hexokinase. The relative amount of ATP contributed by adenylate kinase is dependent upon the ADP concentration. At low ADP concentrations, glucose phosphorylation is supported by oxidative phosphorylation, but as the adenine nucleotide translocator becomes saturated the ATP contributed by adenylate kinase increases due to the higher apparent Km of the enzyme. Under conditions of our standard experiment ([ADP] = 0.5 mM), adenylate kinase provides about 50% of the ATP used by hexokinase in well-coupled mitochondria. In spite of this, externally added ATP supported higher initial rates of hexokinase activity than ADP. Our findings demonstrate that oxidative phosphorylation is not a specific or preferential source of ATP for hexokinase bound to hepatoma mitochondria. The apparent lack of a channeling mechanism for ATP to hexokinase in these mitochondria is discussed.

Studies on beef heart ubiquinol-cytochrome c reductase. Quantification and distribution of the core proteins as determined by radioimmunoassay.

Core proteins I (Mr 50 000) and II (Mr 47 000) were isolated from beef heart ubiquinol-cytochrome c reductase, and radioimmunoassays were developed for both. Immunoreplica experiments show that antisera against each protein react with a single peptide in both isolated Complex III and in mitochondria. Thus, core proteins are not aggregated forms of smaller peptides as suggested for the yeast protein (Jeffrey, A., Power, S. and Palmer, G., Biochem. Biophys. Res. Commun. (1979) 86, 271-277). Core proteins were quantitated in Complex III and in mitochondria using radioimmunoassay. Approx. 2 mol core protein II per mol core protein I were found. A molar ratio of 1 : 2 : 2 : 1 is suggested for core protein I : core protein II : cytochrome b : cytochrome c1. Radioimmunoassay shows that the antibodies react as extensively with Complex III-bound core protein as with the isolated core proteins. In spite of this, the antibodies do not inhibit electron transport in submitochondrial particles or isolated Complex III, and they have no oligomycin- or uncoupler-like effects on submitochondrial particles oxidizing NADH. The combined results from radioimmunoassay and immunoreplica experiments strongly suggest, however, that core proteins are specifically associated with Complex III in the mitochondria, implying a specific role there.

Expression of a new set of glycolytic isozymes in activated human peripheral lymphocytes.

The combination of phorbol 12-myristate 13-acetate (PMA) and concanavalin A induces the expression of a new set of glycolytic isozymes in human peripheral lymphocytes. The induced isozyme for each enzyme tested (hexokinase, phosphofructokinase, enolase, pyruvate kinase and lactate dehydrogenase) is usually the muscle form, which is often associated with rapidly dividing tumor cells. Increases in a specific isozyme can account for the 2-5-fold increase in specific activities of the enzyme induced by Con A plus PMA. Increased specific activities and the appearance of the new isozyme forms both occur relatively late, and are probably associated with the G1 or S phase of the cell cycle.

Inhibition of mitochondrial protein synthesis in regenerating rat liver stimulates mitochondrial transcription.

Partially hepatectomized rats were treated in vivo with thiamphenicol for 3 days to block mitochondrial protein synthesis. Protein synthesis, RNA synthesis and the steady-state levels of individual transcripts were measured in mitochondria in vitro in the absence of thiamphenicol. Incorporation of [35S]methionine and [3H]UTP into protein and RNA, respectively, was increased 2-3-fold in isolated mitochondria from thiamphenicol-treated animals, indicating increased rates of synthesis of both. Electrophoretic analysis of transcripts labelled with [32P]UTP suggests that synthesis of all the transcripts is increased. The steady-state concentrations of mitochondrial transcripts, measured by Northern blotting using nick-translated cloned EcoRI fragments of rat liver mtDNA, were also elevated 2-4-fold in thiamphenicol-treated animals. The data suggest that mitochondrial transcription is under control of a mitochondrial factor which, in turn, is dependent upon mitochondrial protein synthesis.

Effect of phorbol myristate acetate and concanavalin A on the glycolytic enzymes of human peripheral lymphocytes.

The effects of Concanavalin A and the tumor promoting agent, phorbol 12-myristate 13-acetate (PMA), on glycolytic enzymes in human peripheral lymphocytes have been studied. A combination of Concanavalin A plus PMA stimulates DNA and protein synthesis to a significantly greater extent than when each are added individually. PMA and concanavalin A together, but not individually, also increase the levels of the activity of the glycolytic enzymes in peripheral lymphocytes treated for 48 h. The increase in hexokinase activity induced by PMA plus concanavalin A appeared to be due to the expression of the isoenzyme form, hexokinase II. The results suggest that the expression of glycolytic enzymes in stimulated lymphocytes is a late event (perhaps associated with the S phase) which is regulated by a cellular signal system controlled by the combined action of PMA plus concanavalin A.