Background

The WW domain is found in a large number of eukaryotic proteins implicated in a variety of cellular processes. WW domains bind proline-rich protein and peptide ligands, but the protein interaction partners of many WW domain-containing proteins in Saccharomyces cerevisiae are largely unknown.

Results

We used protein microarray technology to generate a protein interaction map for 12 of the 13 WW domains present in proteins of the yeast S. cerevisiae. We observed 587 interactions between these 12 domains and 207 proteins, most of which have not previously been described. We analyzed the representation of functional annotations within the network, identifying enrichments for proteins with peroxisomal localization, as well as for proteins involved in protein turnover and cofactor biosynthesis. We compared orthologs of the interacting proteins to identify conserved motifs known to mediate WW domain interactions, and found substantial evidence for the structural conservation of such binding motifs throughout the yeast lineages. The comparative approach also revealed that several of the WW domain-containing proteins themselves have evolutionarily conserved WW domain binding sites, suggesting a functional role for inter- or intramolecular association between proteins that harbor WW domains. On the basis of these results, we propose a model for the tuning of interactions between WW domains and their protein interaction partners.

Conclusion

Protein microarrays provide an appealing alternative to existing techniques for the construction of protein interaction networks. Here we built a network composed of WW domain-protein interactions that illuminates novel features of WW domain-containing proteins and their protein interaction partners.

The extent and nature of epigenomic changes associated with melanoma progression is poorly understood. Through systematic epigenomic profiling of 35 epigenetic modifications and transcriptomic analysis, we define chromatin state changes associated with melanomagenesis by using a cell phenotypic model of non-tumorigenic and tumorigenic states. Computation of specific chromatin state transitions showed loss of histone acetylations and H3K4me2/3 on regulatory regions proximal to specific cancer-regulatory genes in important melanoma-driving cell signaling pathways. Importantly, such acetylation changes were also observed between benign nevi and malignant melanoma human tissues. Intriguingly, only a small fraction of chromatin state transitions correlated with expected changes in gene expression patterns. Restoration of acetylation levels on deacetylated loci by histone deacetylase (HDAC) inhibitors selectively blocked excessive proliferation in tumorigenic cells and human melanoma cells, suggesting functional roles of observed chromatin state transitions in driving hyperproliferative phenotype. Through these results, we define functionally relevant chromatin states associated with melanoma progression.

Although donation of bone marrow (BM) or peripheral blood stem cells (PBSCs) from children to family members undergoing allogeneic transplantation are well-established procedures, studies detailing levels of pain, symptoms, and long-term recovery are lacking. To address this lack, we prospectively enrolled 294 donors age <18 years at 25 pediatric transplantation centers in North America, assessing them predonation, peridonation, and at 1 month, 6 months, and 1 year postdonation. We noted that 71% of children reported pain and 59% reported other symptoms peridonation, with resolution to 14% and 12% at 1 month postdonation. Both older age (age 13 to 17 years versus younger) and female sex were associated with higher levels of pain peridonation, with the highest rates in older females (57% with grade 2-4 pain and 17% with grade 3-4 pain). Multivariate analyses showed a 4-fold increase in risk for older females compared with males age <13 years (P <.001). At 1 year, 11% of 13- to 17-year-old females reported grade 2-4 pain, compared with 3% of males age 13 to 17 years, 0% of females age <13 years, and 1% of males age <13 years (P = .01). Males and females age 13 to 17 years failed to return to predonation pain levels at 1 year 22% and 23% of the time, respectively, compared with 3% and 10% in males and females age <13 years (P = .002). Our data show that females age 13 to 17 years are at increased risk of grade 2-4 pain at 1 year and >20% of females and males age 13 to 17 years do not return to baseline pain levels by 1 year after BM donation. Studies aimed at decreasing symptoms and improving recovery in older children are warranted.

The development of reduced-intensity approaches for allogeneic hematopoietic cell transplantation has resulted in growing numbers of older related donors (RDs) of peripheral blood stem cells (PBSCs). The effects of age on donation efficacy, toxicity, and long-term recovery in RDs are poorly understood. To address this we analyzed hematologic variables, pain, donation-related symptoms, and recovery in 1211 PBSC RDs aged 18 to 79 enrolled in the Related Donor Safety Study. RDs aged > 60 had a lower median CD34⁺ level before apheresis compared with younger RDs (age > 60, 59 × 10⁶/L; age 41 to 60, 81 × 10⁶/L; age 18 to 40, 121 × 10⁶/L; P < .001). This resulted in older donors undergoing more apheresis procedures (49% versus 30% ≥ 2 collections, P < .001) and higher collection volumes (52% versus 32% > 24 L, P < .001), leading to high percentages of donors aged > 60 with postcollection thrombocytopenia <50 × 10⁹/L (26% and 57% after 2 and 3days of collection, respectively). RDs aged 18 to 40 had a higher risk of grades 2 to 4 pain and symptoms pericollection, but donors over age 40 had more persistent pain at 1, 6, and 12 months (odds ratio [OR], 1.7; P = 0.02) and a higher rate of nonrecovery to predonation levels (OR, 1.7; P = .01). Donors reporting comorbidities increased significantly with age, and those with comorbidities that would have led to deferral by National Marrow Donor Program unrelated donor standards had an increased risk for persistent grades 2 to 4 pain (OR, 2.41; P < .001) and failure to recover to predonation baseline for other symptoms (OR, 2.34; P = .004). This information should be used in counseling RDs regarding risk and can assist in developing practice approaches aimed at improving the RD experience for high-risk individuals.

Unlike unrelated donor registries, transplant centers lack uniform approaches to related donor assessment and deferral. To test whether related donors are at increased risk for donation-related toxicities, we conducted a prospective observational trial of 11,942 related and unrelated donors aged 18-60 years. Bone marrow (BM) was collected at 37 transplant and 78 National Marrow Donor Program centers, and peripheral blood stem cells (PBSC) were collected at 42 transplant and 87 unrelated donor centers in North America. Possible presence of medical comorbidities was verified prior to donation, and standardized pain and toxicity measures were assessed pre-donation, peri-donation, and one year following. Multivariate analyses showed similar experiences for BM collection in related and unrelated donors; however, related stem cell donors had increased risk of moderate [odds ratios (ORs) 1.42; P<0.001] and severe (OR 8.91; P<0.001) pain and toxicities (OR 1.84; P<0.001) with collection. Related stem cell donors were at increased risk of persistent toxicities (OR 1.56; P=0.021) and non-recovery from pain (OR 1.42; P=0.001) at one year. Related donors with more significant comorbidities were at especially high risk for grade 2-4 pain (OR 3.43; P<0.001) and non-recovery from toxicities (OR 3.71; P<0.001) at one year. Related donors with more significant comorbidities were at especially high risk for grade 2-4 pain (OR 3.43; P<0.001) and non-recovery from toxicities (OR 3.71; P<0.001) at one year. Related donors reporting grade ≥2 pain had significant decreases in Health-Related Quality of Life (HR-QoL) scores at one month and one year post donation (P=0.004). In conclusion, related PBSC donors with comorbidities are at increased risk for pain, toxicity, and non-recovery at one year after donation. Risk profiles described in this study should be used for donor education, planning studies to improve the related donor experience, and decisions regarding donor deferral. Registered at clinicaltrials.gov identifier:00948636.

We describe the landscape of genomic alterations in cutaneous melanomas through DNA, RNA, and protein-based analysis of 333 primary and/or metastatic melanomas from 331 patients. We establish a framework for genomic classification into one of four subtypes based on the pattern of the most prevalent significantly mutated genes: mutant BRAF, mutant RAS, mutant NF1, and Triple-WT (wild-type). Integrative analysis reveals enrichment of KIT mutations and focal amplifications and complex structural rearrangements as a feature of the Triple-WT subtype. We found no significant outcome correlation with genomic classification, but samples assigned a transcriptomic subclass enriched for immune gene expression associated with lymphocyte infiltrate on pathology review and high LCK protein expression, a T cell marker, were associated with improved patient survival. This clinicopathological and multi-dimensional analysis suggests that the prognosis of melanoma patients with regional metastases is influenced by tumor stroma immunobiology, offering insights to further personalize therapeutic decision-making.