DNA methylation control of tissue polarity and cellular differentiation in the mammary epithelium

Alterations in gene expression accompany cell-type-specific differentiation. In complex systems where functional differentiation depends on the organization of specific cell types into highly specialized structures (tissue morphogenesis), it is not known how epigenetic mechanisms that control gene expression influence this stepwise differentiation process. We have investigated the effect of DNA methylation, a major epigenetic pathway of gene silencing, on the regulation of mammary acinar differentiation. Our in vitro model of differentiation encompasses human mammary epithelial cells that form polarized and hollow tissue structures (acini) when cultured in the presence of basement membrane components. We found that acinar morphogenesis was accompanied with chromatin remodeling, as shown by alterations in histone 4 acetylation, heterochromatin 1 protein, and histone 3 methylated on lysine 9, and with an increase in expression of MeCP2, a mediator of DNA-methylation-induced gene silencing. DNA hypomethylation induced by treatment with 5-aza-2' deoxycytidine during acinar differentiation essentially prevented the formation of apical tissue polarity. This treatment also induced the expression of CK19, a marker of cells that are in a transitional differentiation stage. These results suggest that DNA methylation is a mechanism by which mammary epithelial differentiation is coordinated both at the tissue and cellular levels.     

Obesity-associated mutations in the melanocortin 4 receptor provide novel insights into its function

Mutations in the Melanocortin 4 receptor are implicated in 1-6% of early onset or severe adult obesity cases. Most of the patients carry heterozygous missense mutations. Arguments for the pathogenicity of these mutations are based on the frequency of rare functionally relevant non-synonymous mutations in severely obese children and adults versus non-obese controls, the segregation of mutations with obesity in the family of the probands (although with incomplete penetrance) and the relevant functional defects described for these mutations. We have developed new assays to study the functional characteristics of these obesity-associated MC4R mutations. Systematic and comparative functional study of over 50 different obesity-associated mutations suggests that multiple functional alterations contribute to their pathogenicity. These studies also lead to new insights into the structure-function relationship of MC4R, provide novel hypotheses for the genetic predisposition to common obesity in humans and allow the development of new molecular tools for studying the physiological role of GPCRs.     

Coffee and Pancreatic Cancer in a Rural California County

In a study of the risk of fatal pancreatic cancer according to intake of regular and decaffeinated coffee, cases (N = 30) and controls (N = 47) were identified from death certificates and matched for age (± 5 years), sex, ethnicity, and date of death (± 5 years). Telephone interviews were completed with survivors of about 80% of both groups. In smokers, the relative risk for high (3 + cups) versus low (<3 cups) intake of regular coffee was 4.3 (P < .05), and high verus low decaffeinated coffee, 5.5 (P < .05). In nonsmokers, neither type of coffee influenced the risk. Mean daily intakes of alcohol and cigarettes were virtually identical in cases and controls, although cases had accumulated nonsignificantly more pack-years. Intakes of regular and decaffeinated coffee were uncorrelated, and the smoking-coffee interaction could not be readily explained by recall bias. If coffee intake increases the risk of pancreatic cancer, the mechanism could depend heavily on smoking.     

Analysis of taxol and related diterpenoids from cell cultures by liquid chromatography—electrospray mass spectrometry

Authentic taxanes (taxol, 10-deacetyltaxol, cephalomannine, 10-deacetylcephalomannine, baccatin III) and extracts from cell cultures derived from various yew tree species have been analyzed by microbore high-performance liquid chromatography (HPLC)—electrospray mass spectrometry (ESMS). All gave excellent positive-ion ES spectra with dominant protonated molecules at low nozzle-to-skimmer bias value (45 V). By increasing the voltage value to 85 V, fragmentation increased and structurally informative spectra were obtained. The fragments found were both of the C-13 side-chain and of the taxane ring, so their analysis gave important information about the taxane structure and any chemical modifications at different positions of the molecule. When tandem MS was used (argon gas, 25 eV collision energy), fragments similar to those obtained from collision-induced dissociation in the source were detected. The cell culture extracts were analyzed by microbore HPLC—ESMS and excellent spectra were obtained on 5–10 ng of separated compounds; even greater selectivity and sensitivity were obtained through use of selected-ion monitoring (SIM). With SIM, 100 pg of all taxanes could readily be detected. In the HPLC—ESMS mode, only 10% of the eluent was mass-analyzed, so 90% would be available for recovery through fraction collecting.     

The influence of kinship and dominance hierarchy on grooming partner choice in free-ranging <Emphasis Type="Italic">Macaca mulatta brevicaudus</Emphasis>

In group-living animals, individuals do not interact uniformly with their conspecifics. Among primates, such heterogeneity in partner choice can be discerned from affiliative grooming patterns. While the preference for selecting close kin as grooming partners is ubiquitous across the primate order, the selection of higher-ranking non-kin individuals as grooming partners is less common. We studied a group of provisioned rhesus macaques (Macaca mulatta brevicaudus) on Hainan Island, China, to examine rank-related benefits of grooming exchanges and the influence of kin relationships. We tested four hypotheses based on Seyfarth’s model: (1) there will be kin preference in grooming relationships; (2) grooming between non-kin individuals will be directed up the dominance rank; (3) grooming between non-kin individuals will reduce aggression from higher-ranking ones; and (4) non-kin individuals will spend more time grooming with adjacent ranked ones. We found that grooming relationships between kin individuals were stronger than those between non-kin individuals. For non-kin relationships, lower-ranking individuals received less aggression from higher-ranking ones through grooming; a benefit they could not derive through grooming exchanges with individuals related by kinship. Individuals spent more time grooming adjacent higher-ranking non-kin individuals and higher-ranking individuals also received more grooming from non-kin individuals. Our results supported Seyfarth’s model for predicting partner choice between non-kin individuals. For relationships between kin individuals, we found results that were not consistent with prediction for the exchanges of aggression and grooming, indicating the importance to control for the influence of kinship in future studies.     

Platelet Releasate Proteome Profiling Reveals a Core Set of Proteins with Low Variance between Healthy Adults

Upon activation, platelets release a powerful cocktail of soluble and vesicular signals, collectively termed the "platelet releasate" (PR). Although several studies have used qualitative/quantitative proteomic approaches to characterize PR; with debated content and significant inter‐individual variability reported, confident, and reliable insights have been hindered. Using label‐free quantitative (LFQ)‐proteomics analysis, a reproducible, quantifiable investigation of the 1U mL^?1 thrombin‐induced PR from 32 healthy adults was conducted. MS proteomics data are available via ProteomeXchange, identifier PXD009310. Of the 894 proteins identified, 277 proteins were quantified across all donors and form a "core" PR. Bioinformatics and further LFQ‐proteomic analysis revealed that the majority (84%) of "core" PR proteins overlapped with the protein composition of human platelet‐derived exosomes. Vesicles in the exosomal‐size range were confirmed in healthy‐human PR and reduced numbers of similar‐sized vesicles were observed in the PR of a mouse model of gray platelet syndrome, known to be deficient in platelet alpha‐granules. Lastly, the variability of proteins in the PR was assessed, and reproducible secretion levels were found across all 32 healthy donors. Taken together, the PR contains valuable soluble and vesicular cargo and has low‐population variance among healthy adults, rendering it a potentially useful platform for diagnostic fingerprinting of platelet‐related disease.     

The role of the frontal ganglion in the control of protein metabolism inLocusta migratoria

Summary 1. After removal of the frontal ganglion, there is an immediate permanent decrease in the blood protein level.2. The cytoplasmic RNA concentration, the blood amino-acid level and the midgut protease level, also decrease after this operation.3. These effects cannot be reversed by feeding as is the case in starved animals.4. The above results provide clear evidence that the frontal ganglion plays a vital part in controlling protein synthesis in the growing locust.

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Die Bedeutung des Frontalganglions für die Kontrolle des Proteinstoffwechsels beiLocusta migratoria

Kurzfassung Am dritten Nymphenstadium vonLocusta migratoria konntenClarke &Langley (1963a) zeigen, daß die Entfernung des Frontalganglions eine völlige Einstellung des Körperwachstums zur Folge hat. Elektrophorese des Blutes operierter Tiere läßt eine starke permanente Abnahme des Proteingehaltes erkennen. Gleichzeitig kommt es zu einer allgemeinen Verringerung des RNS-Gehaltes der Gewebe. Diese ist besonders augenfällig in den aktivsten Geweben, wie etwa der Epidermis und dem Mitteldarmepithel. Weiterhin ergaben papierchromatographische Untersuchungen, daß — im Gegensatz zu der bei Hungertieren angetroffenen Situation — eine Abnahme der Aminosäurekonzentration des Blutes erfolgt, welche ihrerseits wiederum zu einem Zusammenbruch des Wasserhaushaltes führt. Noch nicht abgeschlossene Untersuchungen über die Mitteldarmproteasen deuten auf eine ungenügende Synthese dieser Enzyme in den Epithelzellen der operierten Tiere hin. Es wird eine allgemeine Kontrolle der Proteinsynthese durch das neurosekretorische System postuliert und angenommen, daß dieses System wiederum durch sensorische Impulse kontrolliert wird, welche von den pharyngealen Dehnungsrezeptoren über das Frontalganglion zum Gehirn gelangen.

     

A genome‐wide association study in Caucasian women suggests the involvement of HLA genes in the severity of facial solar lentigines

Solar lentigines are a common feature of sun‐induced skin ageing. Little is known, however, about the genetic factors contributing to their development. In this genome‐wide association study, we aimed to identify genetic loci associated with solar lentigines on the face in 502 middle‐aged French women. Nine SNPs, gathered in two independent blocks on chromosome 6, exhibited a false discovery rate below 25% when looking for associations with the facial lentigine score. The first block, in the 6p22 region, corresponded to intergenic SNPs and also exhibited a significant association with forehead lentigines (P = 1.37 × 10^?8). The second block, within the 6p21 HLA region, was associated with decreased HLA‐C expression according to several eQTL databases. Interestingly, these SNPs were also in high linkage disequilibrium with the HLA‐C*0701 allele (r² = 0.95). We replicated an association recently found by GWAS in the IRF4 gene. Finally, a complementary study on 44 selected candidate SNPs revealed novel associations in the MITF gene. Overall, our results point to several mechanisms involved in the severity of facial lentigines, including HLA/immunity and the melanogenesis pathway.