Studies of the TgHBox4 homeobox gene in Tripneustes gratilla

Thesis (Ph. D.)--University of Hawaii at Manoa, 1994.Includes bibliographical references (leaves 147-154)Microfiche.x, 154 leaves, bound illus. 29 cmThis study further characterized TgHBox4, an Abd-B class homeobox gene expressed as a 4.4kb and a 3.8kb mRNA in early and late embryogenesis, respectively, and as a 3.6kb mRNA in adult tissue of the sea urchin, Tripneustes gratilla. A full length cDNA clone was of 3623 base pairs encoding a 33,312kd protein was selected from an intestinal mRNA library. The cDNA clone was used to select a genomic clone containing a 5' exon of 830bp, which along with an original 3' exon of 2800bp accounted for the full cDNA sequence. Sequence analysis of the cDNA and the 5' genomic clone indicates that the A at the start site of the cDNA may be the transcription start site, with a TATA box 30 nucleotides upstream. The first in frame ATG codon is 210 nucleotides downstream of this transcription start site but the ORF encoding the homeobox protein extends 5' of the start site another 40 nucleotides including an upstream in frame ATG codon. The late embryonic message, which is slightly larger than the intestinal mRNA may be produced by use of an alternate upstream promoter. A TgHBox4 sequence was expressed in bacteria and the protein used as an immunogen. Immunofluorescent studies using the antibodies localized TgHBox4 expression during development. The TgHBox4 proteins appear generally early in embryogenesis and become slowly restricted to the ectoderm cells over the spicule forming primary mesenchyme cells by gastrula stage. As gastrulation proceeds TgHBox4 proteins accumulate in the invaginating gut and became progressively restricted to the region of the digestive tract by pluteus stage. The TgHBox4 bacterially expressed protein was also used in gel shift assays to study it's binding to an enhancer of the late H2B histone gene of S. purpuratus. The anti-TgHBox4 antibodies specifically blocked this binding as well as the binding of the endogenous nuclear factor to the enhancer sequence, indicating that the TgHBox4 gene may regulate late embryonic histone synthesis. It is speculated that the protein product of the larger TgHBox4 message produced in early embryogenesis is involved in the determination of cell patterns which guide mesenchyme cell localization. The protein product of the smaller message produced later in embryogenesis may function to enhance H2B histone gene and other genes expression in dividing cells restricted to the gut in late embryos. The late expression of TgHBox4 in the vegetal plate structures derived from the most posterior parts of the blastula may relate to the posterior expression of the Abd-B class genes in the Hox gene cluster

Case report: value of gene expression profiling in the diagnosis of atypical neuroblastoma

BACKGROUND: Nephroblastoma and neuroblastoma belong to the most common abdominal malignancies in childhood. Similarities in the initial presentation may provide difficulties in distinguishing between these two entities, especially if unusual variations to prevalent patterns of disease manifestation occur. Because of the risk of tumor rupture, European protocols do not require biopsy for diagnosis, which leads to misdiagnosis in some cases. CASE PRESENTATION: We report on a 4½-year-old girl with a renal tumor displaying radiological and laboratory characteristics supporting the diagnosis of nephroblastoma. Imaging studies showed tumor extension into the inferior vena cava and bilateral lung metastases while urine catecholamines and MIBG-scintigraphy were negative. Preoperative chemotherapy with vincristine, actinomycine D and adriamycin according to the SIOP2001/GPOH protocol for the treatment of nephroblastoma was initiated and followed by surgical tumor resection. Histopathology revealed an undifferentiated tumor with expression of neuronal markers, suggestive of neuroblastoma. MYCN amplification could not be detected. DNA-microarray analysis was performed using Affymetrix genechip human genome U133 plus 2.0 and artificial neural network analysis. Results were confirmed by multiplex RT-PCR. RESULTS: Principal component analysis using 84 genes showed that the patient sample was clearly clustering with neuroblastoma tumors. This was confirmed by hierarchical clustering of the multiplex RT-PCR data. The patient underwent treatment for high-risk neuroblastoma comprising chemotherapy including cisplatin, etoposide, vindesine, dacarbacine, ifosfamide, vincristine, adriamycine and autologous stem cell transplantation followed by maintenance therapy with 13-cis retinoic acid (GPOH NB2004 High Risk Trial Protocol) and is in complete long-term remission. CONCLUSION: The use of gene expression profiling in an individual patient strongly contributed to clarification in a diagnostic dilemma which finally led to a change of diagnosis from nephroblastoma to neuroblastoma. This case underlines the importance of gene-expression profiling in the correct diagnosis of childhood neoplasms with atypical presentation to ensure that adequate treatment regimens can be applied

Diagnosis of the Small Round Blue Cell Tumors Using Multiplex Polymerase Chain Reaction

The small round blue cell tumors of childhood, which include neuroblastoma, rhabdomyosarcoma, non-Hodgkin’s lymphoma, and the Ewing’s family of tumors, are so called because of their similar appearance on routine histology. Using cDNA microarray gene expression profiles and artificial neural networks (ANNs), we previously identified 93 genes capable of diagnosing these cancers. Using a subset of these, together with some additional genes (total 39), we developed a multiplex polymerase chain reaction (PCR) assay to diagnose these cancer types. Blinded testing of 96 new samples (26 Ewing’s family of tumors, 29 rhabdomyosarcomas, 24 neuroblastomas, and 17 lymphomas) using ANNs in a complete leave-one-out analysis demonstrated that all except one sample were accurately diagnosed as their respective category. Moreover, using an ANN-based gene minimization strategy in a separate analysis, we found that the top 31 genes could correctly diagnose all 96 tumors. Our results suggest that this molecular test based on a multiplex PCR reaction may assist the physician in the rapid confirmation of the diagnosis of these cancers

Exploring Molecular Pathways of Triple-Negative Breast Cancer

Triple-negative breast cancer (TNBC) is an aggressive breast cancer subtype with a high rate of proliferation and metastasis, as well as poor prognosis for advanced-stage disease. Although TNBC was previously classified together with basal-like and BRCA1/2-related breast cancers, genomic profiling now shows that there is incomplete overlap, with important distinctions associated with each subtype. The biology of TNBC is still poorly understood; therefore, to define the relative contributions of major cellular pathways in TNBC, we have studied its molecular signature based on analysis of gene expression. Comparisons were then made with normal breast tissue. Our results suggest the existence of molecular networks in TNBC, characterized by explicit alterations in the cell cycle, DNA repair, nucleotide synthesis, metabolic pathways, NF-κB signaling, inflammatory response, and angiogenesis. Moreover, we also characterized TNBC as a cancer of mixed phenotypes, suggesting that TNBC extends beyond the basal-like molecular signature and may constitute an independent subtype of breast cancer. The data provide a new insight into the biology of TNBC

MOESM2 of Case report: value of gene expression profiling in the diagnosis of atypical neuroblastoma

Additional file 2: Figure S1. Timeline according to CARE Guidelines for case reports

Involvement of the MADS-Box Gene ZMM4 in Floral Induction and Inflorescence Development in Maize1[W][OA]

The switch from vegetative to reproductive growth is marked by the termination of vegetative development and the adoption of floral identity by the shoot apical meristem (SAM). This process is called the floral transition. To elucidate the molecular determinants involved in this process, we performed genome-wide RNA expression profiling on maize (Zea mays) shoot apices at vegetative and early reproductive stages using massively parallel signature sequencing technology. Profiling revealed significant up-regulation of two maize MADS-box (ZMM) genes, ZMM4 and ZMM15, after the floral transition. ZMM4 and ZMM15 map to duplicated regions on chromosomes 1 and 5 and are linked to neighboring MADS-box genes ZMM24 and ZMM31, respectively. This gene order is syntenic with the vernalization1 locus responsible for floral induction in winter wheat (Triticum monococcum) and similar loci in other cereals. Analyses of temporal and spatial expression patterns indicated that the duplicated pairs ZMM4-ZMM24 and ZMM15-ZMM31 are coordinately activated after the floral transition in early developing inflorescences. More detailed analyses revealed ZMM4 expression initiates in leaf primordia of vegetative shoot apices and later increases within elongating meristems acquiring inflorescence identity. Expression analysis in late flowering mutants positioned all four genes downstream of the floral activators indeterminate1 (id1) and delayed flowering1 (dlf1). Overexpression of ZMM4 leads to early flowering in transgenic maize and suppresses the late flowering phenotype of both the id1 and dlf1 mutations. Our results suggest ZMM4 may play roles in both floral induction and inflorescence development