BackgroundCardiac contractility is regulated by dynamic phosphorylation of sarcomeric proteins by kinases such as cAMP-activated protein kinase A (PKA). Efficient phosphorylation requires that PKA be anchored close to its targets by A-kinase anchoring proteins (AKAPs). Cardiac Myosin Binding Protein-C (cMyBPC) and cardiac troponin I (cTNI) are hypertrophic cardiomyopathy (HCM)-causing sarcomeric proteins which regulate contractility in response to PKA phosphorylation.ResultsDuring a yeast 2-hybrid (Y2H) library screen using a trisphosphorylation mimic of the C1-C2 region of cMyBPC, we identified isoform 4 of myomegalin (MMGL) as an interactor of this N-terminal cMyBPC region. As MMGL has previously been shown to interact with phosphodiesterase 4D, we speculated that it may be a PKA-anchoring protein (AKAP).To investigate this possibility, we assessed the ability of MMGL isoform 4 to interact with PKA regulatory subunits R1A and R2A using Y2H-based direct protein-protein interaction assays. Additionally, to further elucidate the function of MMGL, we used it as bait to screen a cardiac cDNA library. Other PKA targets, viz. CARP, COMMD4, ENO1, ENO3 and cTNI were identified as putative interactors, with cTNI being the most frequent interactor.We further assessed and confirmed these interactions by fluorescent 3D-co-localization in differentiated H9C2 cells as well as by in vivo co-immunoprecipitation. We also showed that quantitatively more interaction occurs between MMGL and cTNI under β-adrenergic stress. Moreover, siRNA-mediated knockdown of MMGL leads to reduction of cMyBPC levels under conditions of adrenergic stress, indicating that MMGL-assisted phosphorylation is requisite for protection of cMyBPC against proteolytic cleavage.ConclusionsThis study ascribes a novel function to MMGL isoform 4: it meets all criteria for classification as an AKAP, and we show that is involved in the phosphorylation of cMyBPC as well as cTNI, hence MMGL is an important regulator of cardiac contractility. This has further implications for understanding the patho-aetiology of HCM-causing mutations in the genes encoding cMyBPC and cTNI, and raises the question of whether MMGL might itself be considered a candidate HCM-causing or modifying factor.

Plasmablastic lymphoma (PBL) is reported rarely in children. To date, 10 cases are documented in the English-language literature. This study, based on 13 biopsies from 11 HIV-positive children (9 males, 2 females), documents the clinicopathologic features of PBL. The CD4 count ranged from 9 to 800 cells/mm(3). All biopsies demonstrated exclusive plasmablastic morphology; CD20 immunonegativity; and VS38c, EMA, CD31, MUM-1, CD45, and CD79a immunopositivity. B-cell monoclonality was confirmed in all biopsies. Of 3 biopsies subjected to FISH investigation, 2 had a t(8,14) translocation. Nine patients with follow-up details were treated exclusively with HAART (highly active antiretroviral therapy) or with combinations of HAART, chemotherapy, and radiotherapy. Seven patients died. PBL histomorphology, disease stage, and treatment modalities employed were not predictive of outcome. The survival of 2 stage 4 patients for 3 and 8 years each, managed on HAART, chemotherapy, and radiotherapy, however, may justify a role for combined therapeutic modalities for PBL.

Aims: To investigate the incidence of genetic aberrations in the DNA repair genes in a cohort of oesophageal cancers. Methods: One hundred oesophagectomy samples of squamous cell carcinoma were studied. Normal and tumour DNA were isolated using a standard phenol/chloroform extraction procedure. Six recommended microsatellite loci with high informativity were analysed. The following markers were used: D2S123 (2p), D3S659 (3p), D3S1255 (3p), Bat 25 (4q), Bat 26 (2p), and Bat 40 (1p). The results were analysed using software attached to an automated DNA sequencer. The molecular data were then correlated with clinicopathological parameters. Results: The incidence of microsatellite instability and loss of heterozygosity was very low. There was no significant correlation between the clinicopathological and molecular data. However, D2S123 genetic abnormalities were seen more frequently in both moderately and well differentiated tumours than in poorly differentiated tumours (p = 0.033). Follow up data were available for only 67 of the 100 patients. Fifty patients were alive and 17 patients had died. Conclusion: Low frequencies of genetic aberrations in these mismatch repair loci are found in squamous carcinomas of the oesophagus from a high incidence area in South Africa. O esophageal cancer is the most common malignancy in black South African men, accounting for 14.3% of all cancers in South Africa.1 Over and above the various environmental factors that impact on the development of cancer in general, great advances have been made during the past decade in our understanding of the molecular mechanisms that occur in the transition from normal mucosa, through dysplasia, and finally to carcinoma.2-4 Several studies have revealed that an accumulation of genetic alterations gives rise to cancer. Furthermore, microsatellite instability (MSI) and loss of heterozygosity (LOH) at highly informative genetic loci at which tumour suppressor genes and oncogenes are located are thought to play a role in carcinogenesis. [5][6][7][8][9][10] Genes investigated in recent studies include p53, Rb, DCC, APC, and MCC. 11 We recently showed that the occurrence of MSI in South African squamous carcinomas of the oesophagus was similar to other high incidence areas. ''The mismatch repair genes play an important role in ensuring that errors occurring during DNA synthesis are corrected'' However, very little is known about the status of the mismatch repair genes and their impact on oesophageal cancer. The mismatch repair genes play an important role in ensuring that errors occurring during DNA synthesis are corrected. These genes were initially identified in bacterial systems and mutations in DNA mismatch repair genes have been associated with hereditary non-polyposis colorectal cancer.12-14 Five human DNA mismatch repair genes have been identified, which when mutated cause susceptibility to both sporadic and hereditary non-polyposis colorectal cancer. Mutational inactivation of both copies of a mismatch repair gene results in a profound repair de...

Aims:To determine whether microsatellite mutations of the adenomatous polyposis coli (APC) gene have pathological or prognostic significance in nephroblastomas and to correlate APC alterations with β catenin immunoexpression.Methods:One hundred nephroblastomas were analysed, 83 of which received preoperative chemotherapy. Normal and tumour DNA was isolated using standard proteinase K digestion and phenol/chloroform extraction from paraffin wax embedded tissue. Polymerase chain reaction using four APC microsatellite markers—D5S210, D5S299, D5S82, and D5S346—was performed and the products analysed. Immunohistochemistry was performed using the LSAB kit with diaminobenzidine as chromogen. Results were correlated with clinicopathological data using the χ2test.Results:Allelic imbalance/loss of heterozygosity was more frequent than microsatellite instability, with 30% of cases showing allelic imbalance/ loss of heterozygosity and 16% showing microsatellite instability. Although there was a significant correlation between the results for individual markers and the clinicopathological data, the overall results do not support a prognostic role for APC in nephroblastoma. Expression of β catenin was seen in 93% of cases. Staining was predominantly membranous, with epithelium, blastema, and stroma being immunoreactive. Cytoplasmic redistribution was seen in 58% of cases, but no nuclear staining was detected. No significant associations between β catenin expression and the clinicopathological parameters were found. Kaplan–Meier survival plots showed that patients with loss of membranous staining and pronounced cytoplasmic staining (score, 3) had a significantly shorter survival (p = 0.04; median survival, 5.87 months).Conclusion:Microsatellite analysis of APC and immunoexpression of β catenin did not provide significant pathological or prognostic information in this cohort of nephroblastomas.

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