GTP analogues cause release of the alpha subunit of the GTP binding protein, GO, from the plasma membrane of NG108-15 cells

Incubation of membranes of neuroblastoma x glioma hybrid, NG108-15 cells with GDP beta S followed by immunoblotting of resolved membrane and supernatant fractions with specific anti-peptide antisera showed essentially all of the alpha subunit of Go to be associated with the membrane. Similar experiments with poorly hydrolyzed analogues of GTP caused release of a significant fraction (some 50% within 60 minutes) of Go alpha into the supernatant. This was not mimicked by analogues of ATP. Antisera directed against peptides corresponding to the extreme N and C-termini of GO alpha demonstrated that the released polypeptide was not proteolytically clipped. These experiments show that the alpha subunit of GO need not be invariably bound to the plasma membrane and that guanine nucleotide activation can release the alpha subunit of GO from its site of membrane attachment.     

Impact of variability among surgeons on postoperative morbidity and mortality and ultimate survival.

OBJECTIVE--To assess the differences among surgeons in postoperative complications, postoperative mortality, and survival in patients undergoing surgery for colorectal cancer. DESIGN--Prospective study of patients with colorectal cancer managed by one of 13 consultant surgeons, none of whom had a special interest in colorectal surgery. SETTING--Royal Infirmary, Glasgow. PATIENTS--645 sequential patients with colorectal cancer presenting over the six years from 1974 to 1979. MAIN OUTCOME MEASURES--Postoperative complications, postoperative mortality (within 30 days), and survival (up to 10 years); predictive factors for postoperative mortality and survival; and relative hazard rate ratios for individual surgeons. RESULTS--The proportion of patients undergoing apparently curative resection varied among surgeons from 40% to 76%; overall postoperative mortality varied from 8% to 30%. After curative resection postoperative mortality varied from 0% to 20%, local recurrence from 0% to 21%, and the rate of anastomotic leak from 0% to 25%. Survival at 10 years in patients who underwent curative resection varied from 20% to 63%, two year survival in those who underwent palliative resection varied from 7% to 32%, and median survival in those who underwent palliative diversion varied from one to eight months. The hazard rate ratios among individual surgeons, taking into account the identified risk factors, varied from 0.56 to 2.03, from 0.17 to 1.92, and from 0.57 to 1.50 for curative resection, palliative resection, and palliative diversion, respectively. CONCLUSION--There were significant variations in patient outcome among surgeons after surgery for colorectal cancer; such differences compromise survival. A considerable improvement in overall survival might be achieved if such surgery were undertaken by surgeons with a special interest in colorectal surgery or surgical oncology.     

A PATO-compliant zebrafish screening database (MODB): management of morpholino knockdown screen information

Background  

The zebrafish is a powerful model vertebrate amenable to high throughput in vivo genetic analyses. Examples include reverse genetic screens using morpholino knockdown, expression-based screening using enhancer trapping and forward genetic screening using transposon insertional mutagenesis. We have created a database to facilitate web-based distribution of data from such genetic studies.     

The role of CD4+ T cell help in cancer immunity and the formulation of novel cancer vaccines

Recent years have seen the unprecedented surge of interest in the role of CD4+ T cells and the role they play in the development of the immune response. In this symposium review, we examine the evidence for this and discuss their functions, particularly in respect to the cancer immunology, including CD4+CD25+ cells (Treg).This article is a symposium paper from the conference “Progress in Vaccination against Cancer 2005 (PIVAC 5)”, held in Athens, Greece, on 20–21 September 2005.Deepak P. Assudani, Roger B.V. Horton and Morgan G. Mathieu are joint first authors and have contributed equally to this paper.     

Interrelations between ceruloplasmin and Fe status during human pregnancy

It is well established that Fe and ceruloplasmin interact in animals and in in vitro models. However, Fe-mediated regulation of ceruloplasmin has never been investigated in humans. In an observational study, 53 pregnant women aged 19–39 yr (29.8±0.7 yr, mean ± SEM) were recruited at the Aberdeen Antenatal Clinic, Aberdeen Maternity Hospital, UK. All requirements for local ethical committees were followed. Venous blood samples were taken from each woman at 34 wk gestation for measurement of Fe status and ceruloplasmin. Various parameters were used to test for Fe status. The most sensitive one appeared to be soluble transferrin receptor, which increased with parity. In the population studied, there was no relationship between hemoglobin or ferritin and serum ceruloplasmin. However, using soluble transferrin receptor (sTfR) levels, we were able to demonstrate an inverse linear relationship (r=0.37, p=0.021, n=41) between Fe status and ceruloplasmin. Fe supplementation, number of previous pregnancies, and smoking habits did not affect this relationship. Our data support in vitro results showing regulation of ceruloplasmin by Fe and also suggest that the interactions between Fe and ceruloplasmin should be considered when Fe supplementation is given.     

The helicase HAGE expressed by malignant melanoma-initiating cells is required for tumor cell proliferation in vivo

Malignant melanoma-initiating cells (MMIC) are a subpopulation of cells responsible for melanoma tumor growth and progression. They are defined by the expression of the ATP-binding cassette (ABC) subfamily B member 5 (ABCB5). Here, we identified a critical role for the DEAD-box helicase antigen (HAGE) in ABCB5+ MMIC-dependent tumorigenesis and show that HAGE-specific inactivation inhibits melanoma tumor growth mediated by this tumor-initiating population. Knockdown of HAGE led to a significant decrease in RAS protein expression with a concomitant decrease in activation of the AKT and ERK signaling pathways implicated to play an important role in melanoma progression. To confirm that the reduction in NRAS (Neuroblastoma RAS) expression was dependent on the HAGE helicase activity, we showed that NRAS, effectively silenced by siRNA, could be rescued by reintroduction of HAGE in cells lacking HAGE. Furthermore, we provide a mechanism by which HAGE promotes NRAS unwinding in vitro. We also observed using tumor transplantation in Non-obese diabetic/severe combined immunodeficiency mice that the HAGE knockdown in a ABCB5+ melanoma cell line displayed a significant decrease in tumor growth and compared with the control. Our results suggest that the helicase HAGE is required for ABCB5+ MMIC-dependent tumor growth through promoting RAS protein expression and that cancer therapies targeting HAGE helicase may have broad applications for treating malignant melanoma and potentially other cancer types.