ASU Scholarship Showcase

Human protein diversity arises as a result of alternative splicing, single nucleotide polymorphisms (SNPs) and posttranslational modifications. Because of these processes, each protein can exists as multiple variants in vivo. Tailored strategies are needed to study these protein variants and understand their role in health and disease. In this work we utilized quantitative mass spectrometric immunoassays to determine the protein variants concentration of beta-2-microglobulin, cystatin C, retinol binding protein, and transthyretin, in a population of 500 healthy individuals. Additionally, we determined the longitudinal concentration changes for the protein variants from four individuals over a 6 month period. Along with the native forms of the four proteins, 13 posttranslationally modified variants and 7 SNP-derived variants were detected and their concentration determined. Correlations of the variants concentration with geographical origin, gender, and age of the individuals were also examined. This work represents an important step toward building a catalog of protein variants concentrations and examining their longitudinal changes.

Background: Meiotic recombination has traditionally been explained based on the structural requirement to stabilize homologous chromosome pairs to ensure their proper meiotic segregation. Competing hypotheses seek to explain the emerging findings of significant heterogeneity in recombination rates within and between genomes, but intraspecific comparisons of genome-wide recombination patterns are rare. The honey bee (Apis mellifera) exhibits the highest rate of genomic recombination among multicellular animals with about five cross-over events per chromatid.

Results: Here, we present a comparative analysis of recombination rates across eight genetic linkage maps of the honey bee genome to investigate which genomic sequence features are correlated with recombination rate and with its variation across the eight data sets, ranging in average marker spacing ranging from 1 Mbp to 120 kbp. Overall, we found that GC content explained best the variation in local recombination rate along chromosomes at the analyzed 100 kbp scale. In contrast, variation among the different maps was correlated to the abundance of microsatellites and several specific tri- and tetra-nucleotides.

Conclusions: The combined evidence from eight medium-scale recombination maps of the honey bee genome suggests that recombination rate variation in this highly recombining genome might be due to the DNA configuration instead of distinct sequence motifs. However, more fine-scale analyses are needed. The empirical basis of eight differing genetic maps allowed for robust conclusions about the correlates of the local recombination rates and enabled the study of the relation between DNA features and variability in local recombination rates, which is particularly relevant in the honey bee genome with its exceptionally high recombination rate.

Proteins can exist as multiple proteoforms in vivo, as a result of alternative splicing and single-nucleotide polymorphisms (SNPs), as well as posttranslational processing. To address their clinical significance in a context of diagnostic information, proteoforms require a more in-depth analysis. Mass spectrometric immunoassays (MSIA) have been devised for studying structural diversity in human proteins. MSIA enables protein profiling in a simple and high-throughput manner, by combining the selectivity of targeted immunoassays, with the specificity of mass spectrometric detection. MSIA has been used for qualitative and quantitative analysis of single and multiple proteoforms, distinguishing between normal fluctuations and changes related to clinical conditions. This mini review offers an overview of the development and application of mass spectrometric immunoassays for clinical and population proteomics studies. Provided are examples of some recent developments, and also discussed are the trends and challenges in mass spectrometry-based immunoassays for the next-phase of clinical applications.

Honeybee workers are essentially sterile female helpers that make up the majority of individuals in a colony. Workers display a marked change in physiology when they transition from in-nest tasks to foraging. Recent technological advances have made it possible to unravel the metabolic modifications associated with this transition. Previous studies have revealed extensive remodeling of brain, thorax, and hypopharyngeal gland biochemistry. However, data on changes in the abdomen is scarce. To narrow this gap we investigated the proteomic composition of abdominal tissue in the days typically preceding the onset of foraging in honeybee workers.

In order to get a broader representation of possible protein dynamics, we used workers of two genotypes with differences in the age at which they initiate foraging. This approach was combined with RNA interference-mediated downregulation of an insulin/insulin-like signaling component that is central to foraging behavior, the insulin receptor substrate (irs), and with measurements of glucose and lipid levels.
Our data provide new insight into the molecular underpinnings of phenotypic plasticity in the honeybee, invoke parallels with vertebrate metabolism, and support an integrated and irs-dependent association of carbohydrate and lipid metabolism with the transition from in-nest tasks to foraging.

Background: Omnivorous diets are high in arachidonic acid (AA) compared to vegetarian diets. Research shows that high intakes of AA promote changes in brain that can disturb mood. Omnivores who eat fish regularly increase their intakes of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), fats that oppose the negative effects of AA in vivo. In a recent cross-sectional study, omnivores reported significantly worse mood than vegetarians despite higher intakes of EPA and DHA. This study investigated the impact of restricting meat, fish, and poultry on mood.

Findings: Thirty-nine omnivores were randomly assigned to a control group consuming meat, fish, and poultry daily (OMN); a group consuming fish 3-4 times weekly but avoiding meat and poultry (FISH), or a vegetarian group avoiding meat, fish, and poultry (VEG). At baseline and after two weeks, participants completed a food frequency questionnaire, the Profile of Mood States questionnaire and the Depression Anxiety and Stress Scales. After the diet intervention, VEG participants reduced their EPA, DHA, and AA intakes, while FISH participants increased their EPA and DHA intakes. Mood scores were unchanged for OMN or FISH participants, but several mood scores for VEG participants improved significantly after two weeks.

Conclusions: Restricting meat, fish, and poultry improved some domains of short-term mood state in modern omnivores. To our knowledge, this is the first trial to examine the impact of restricting meat, fish, and poultry on mood state in omnivores.

Background: Mutual policing is an important mechanism for reducing conflict in cooperative groups. In societies of ants, bees, and wasps, mutual policing of worker reproduction can evolve when workers are more closely related to the queen's sons than to the sons of workers or when the costs of worker reproduction lower the inclusive fitness of workers. During colony growth, relatedness within the colony remains the same, but the costs of worker reproduction may change. The costs of worker reproduction are predicted to be greatest in incipient colonies. If the costs associated with worker reproduction outweigh the individual direct benefits to workers, policing mechanisms as found in larger colonies may be absent in incipient colonies.

Results: We investigated policing behavior across colony growth in the ant 'Camponotus floridanus.' In large colonies of this species, worker reproduction is policed by the destruction of worker-laid eggs. We found workers from incipient colonies do not exhibit policing behavior, and instead tolerate all conspecific eggs. The change in policing behavior is consistent with changes in egg surface hydrocarbons, which provide the informational basis for policing; eggs laid by queens from incipient colonies lack the characteristic hydrocarbons on the surface of eggs laid by queens from large colonies, making them chemically indistinguishable from worker-laid eggs. We also tested the response to fertility information in the context of queen tolerance. Workers from incipient colonies attacked foreign queens from large colonies; whereas workers from large colonies tolerated such queens. Workers from both incipient and large colonies attacked foreign queens from incipient colonies.

Conclusions: Our results provide novel insights into the regulation of worker reproduction in social insects at both the proximate and ultimate levels. At the proximate level, our results show that mechanisms of social regulation, such as the response to fertility signals, change dramatically over a colony's life cycle. At the ultimate level, our results emphasize the importance of factors besides relatedness in predicting the level of conflict within a colony. Our results also suggest policing may not be an important regulatory force at every stage of colony development. Changes relating to the life cycle of the colony are sufficient to account for major differences in social regulation in an insect colony. Mechanisms of conflict mediation observed in one phase of a social group's development cannot be generalized to all stages.

Introduction: Apolipoprotein C-III (apoC-III) regulates triglyceride (TG) metabolism. In plasma, apoC-III exists in non-sialylated (apoC-III_0a without glycosylation and apoC-III[subscript 0b] with glycosylation), monosialylated (apoC-III₁) or disialylated (apoC-III₂) proteoforms. Our aim was to clarify the relationship between apoC-III sialylation proteoforms with fasting plasma TG concentrations.

Methods: In 204 non-diabetic adolescent participants, the relative abundance of apoC-III plasma proteoforms was measured using mass spectrometric immunoassay.

Results: Compared with the healthy weight subgroup (n = 16), the ratios of apoC-III_0a, apoC-III_0b, and apoC-III₁ to apoC-III₂ were significantly greater in overweight (n = 33) and obese participants (n = 155). These ratios were positively correlated with BMI z-scores and negatively correlated with measures of insulin sensitivity (S[subscript i]). The relationship of apoC-III₁ / apoC-III₂ with S_i persisted after adjusting for BMI (p = 0.02). Fasting TG was correlated with the ratio of apoC-III_0a / apoC-III₂ (r = 0.47, p<0.001), apoC-III_0b / apoC-III₂ (r = 0.41, p<0.001), apoC-III₁ / apoC-III₂ (r = 0.43, p<0.001). By examining apoC-III concentrations, the association of apoC-III proteoforms with TG was driven by apoC-III_0a (r = 0.57, p<0.001), apoC-III_0b (r = 0.56. p<0.001) and apoC-III₁ (r = 0.67, p<0.001), but not apoC-III₂ (r = 0.006, p = 0.9) concentrations, indicating that apoC-III relationship with plasma TG differed in apoC-III₂ compared with the other proteoforms.

Conclusion: We conclude that apoC-III_0a, apoC-III_0b, and apoC-III₁, but not apoC-III₂ appear to be under metabolic control and associate with fasting plasma TG. Measurement of apoC-III proteoforms can offer insights into the biology of TG metabolism in obesity.

Background: The physical health status of vegetarians has been extensively reported, but there is limited research regarding the mental health status of vegetarians, particularly with regard to mood. Vegetarian diets exclude fish, the major dietary source of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), critical regulators of brain cell structure and function. Omnivorous diets low in EPA and DHA are linked to impaired mood states in observational and experimental studies.

Methods: We examined associations between mood state and polyunsaturated fatty acid intake as a result of adherence to a vegetarian or omnivorous diet in a cross-sectional study of 138 healthy Seventh Day Adventist men and women residing in the Southwest. Participants completed a quantitative food frequency questionnaire, Depression Anxiety Stress Scale (DASS), and Profile of Mood States (POMS) questionnaires.

Results: Vegetarians (VEG:n = 60) reported significantly less negative emotion than omnivores (OMN:n = 78) as measured by both mean total DASS and POMS scores (8.32 ± 0.88 vs 17.51 ± 1.88, p = .000 and 0.10 ± 1.99 vs 15.33 ± 3.10, p = .007, respectively). VEG reported significantly lower mean intakes of EPA (p < .001), DHA (p < .001), as well as the omega-6 fatty acid, arachidonic acid (AA; p < .001), and reported higher mean intakes of shorter-chain α-linolenic acid (p < .001) and linoleic acid (p < .001) than OMN. Mean total DASS and POMS scores were positively related to mean intakes of EPA (p < 0.05), DHA (p < 0.05), and AA (p < 0.05), and inversely related to intakes of ALA (p < 0.05), and LA (p < 0.05), indicating that participants with low intakes of EPA, DHA, and AA and high intakes of ALA and LA had better mood.

Conclusions: The vegetarian diet profile does not appear to adversely affect mood despite low intake of long-chain omega-3 fatty acids.

Background: Peanut consumption favorably influences satiety. This study examined the acute effect of peanut versus grain bar preloads on postmeal satiety and glycemia in healthy adults and the long-term effect of these meal preloads on body mass in healthy overweight adults.

Methods: In the acute crossover trial (n = 15; 28.4 ± 2.9 y; 23.1 ± 0.9 kg/m²), the preload (isoenergetic peanut or grain bar with water, or water alone) was followed after 60 min with ingestion of a standardized glycemic test meal. Satiety and blood glucose were assessed immediately prior to the preload and to the test meal, and for two hours postmeal at 30-min intervals. In the parallel-arm, randomized trial (n = 44; 40.5 ± 1.6 y, 31.8 ± 0.9 kg/m²), the peanut or grain bar preload was consumed one hour prior to the evening meal for eight weeks. Body mass was measured at 2-week intervals, and secondary endpoints included blood hemoglobin A1c and energy intake as assessed by 3-d diet records collected at pre-trial and trial weeks 1 and 8.

Results: Satiety was elevated in the postprandial period following grain bar ingestion in comparison to peanut or water ingestion (p = 0.001, repeated-measures ANOVA). Blood glucose was elevated one hour after ingestion of the grain bar as compared to the peanut or water treatments; yet, total glycemia did not vary between treatments in the two hour postprandial period. In the 8-week trial, body mass was reduced for the grain bar versus peanut groups after eight weeks (−1.3 ± 0.4 kg versus −0.2 ± 0.3 kg, p = 0.033, analysis of covariance). Energy intake was reduced by 458 kcal/d in the first week of the trial for the grain bar group as compared to the peanut group (p = 0.118). Hemoglobin A1c changed significantly between groups during the trial (−0.25 ± 0.07% and −0.18 ± 0.12% for the grain bar and peanut groups respectively, p = 0.001).

Conclusions: Compared to an isoenergetic peanut preload, consumption of a grain bar preload one hour prior to a standardized meal significantly raised postmeal satiety. Moreover, consumption of the grain bar prior to the evening meal was associated with significant weight loss over time suggesting that glycemic carbohydrate ingestion prior to meals may be a weight management strategy.

The impetus for discovery and evaluation of protein biomarkers has been accelerated by recent development of advanced technologies for rapid and broad proteome analyses. Mass spectrometry (MS)-based protein assays hold great potential for in vitro biomarker studies. Described here is the development of a multiplex mass spectrometric immunoassay (MSIA) for quantification of apolipoprotein C-I (apoC-I), apolipoprotein C-II (apoC-II), apolipoprotein C-III (apoC-III) and their proteoforms. The multiplex MSIA assay was fast (∼40 min) and high-throughput (96 samples at a time). The assay was applied to a small cohort of human plasma samples, revealing the existence of multiple proteoforms for each apolipoprotein C. The quantitative aspect of the assay enabled determination of the concentration for each proteoform individually. Low-abundance proteoforms, such as fucosylated apoC-III, were detected in less than 20% of the samples. The distribution of apoC-III proteoforms varied among samples with similar total apoC-III concentrations. The multiplex analysis of the three apolipoproteins C and their proteoforms using quantitative MSIA represents a significant step forward toward better understanding of their physiological roles in health and disease.