PubMed Health⌕ Search

Biomedical subjects

T Iftner

Publications and source records attributed to T Iftner.

30 records · Page 2Linked to original sources
PubMed1994

Changes in RNA expression pattern during the malignant progression of cottontail rabbit papillomavirus-induced tumors in rabbits.

Cottontail rabbit papillomavirus induces strictly epithelial tumors in both cottontail and domestic rabbits. A high proportion of the initial benign papillomas progress within 8 to 14 months to invasive carcinomas. With the help of mRNA-specific riboprobes for E6, E7, E1, E2, L1 and L2, we investigated by in situ hybridization the RNA expression pattern of cottontail rabbit papillomavirus in tissue sections of biopsies from different stages of tumor development. Common features of all lesions were high levels of E6 and E7 mRNAs and low levels of E1 and E2 mRNAs. In agreement with earlier reports, there was no evidence for a major mRNA class equivalent to the prominent E1-E4 RNA of human papillomavirus types 6/11 and 16. In cottontail rabbit papillomas, high levels of E6 and E7 mRNAs were present in the upper differentiated epithelial layers. These layers also contained most of the E1 and E2 mRNAs and the viral DNA. In contrast, papillomas of domestic rabbits revealed the opposite differentiation-dependent expression pattern for the E6 and E7 mRNAs; there were strong signals in the basal layers, and these declined with increased differentiation. Transcripts encoding the L1 mRNA were detected only in a few isolated cells of the granular layer. There was no difference between the amounts of E6, E7, E1, and E2 mRNAs present in highly dysplastic tissue and those present in adjacent normal papillomatous epithelium within a progressing papilloma. However, late transcripts and viral DNA detectable only in the upper layers of the papilloma were present throughout the thickness of the dysplastic tissue, indicating a newly acquired permissiveness of the dysplastic cells for viral DNA replication and late transcription. Carcinomas in general had the same expression patterns for E6, E7, and E1 but were dissimilar in the levels of expression of E2 and late transcripts.

Animals↗
PubMed1993

The predominant mRNA class in HPV16-infected genital neoplasias does not encode the E6 or the E7 protein.

Human papillomavirus (HPV) type 16 is strongly implicated in the development of progressive neoplasias of the uterine cervix. Its oncogenic potential is decisively determined by the activity of the early gene products E6 and E7. To look for changes in the expression of these genes during tumour progression we cloned subgenomic fragments of HPV16 into RNA expression vectors, which allowed the generation of 35S-labelled riboprobes specific for distinct mRNA classes. Four constructs were made to differentiate between transcripts starting upstream of the E6 ORF or the E1 ORF, and one probe was specific for unspliced E6/E7 region transcripts. Five other constructs were used to identify transcripts covering the E1, E2, E4, L1 and L2 regions. With the help of these constructs, we analyzed by in situ hybridization 2 low-grade intraepithelial neoplasias of the vulva, 1 high-grade neoplasia of the cervix as well as 4 vulvar and 3 cervical carcinomas. Transcripts from the E1, E2, E4, L1 and L2 region that were consistently detected in the differentiated layers of benign lesions were variably expressed in precancers and carcinomas. None of the investigated cases revealed detectable amounts of unspliced E6/E7 transcripts with a coding potential for a full-length E6 protein. In benign lesions, the E7 transcripts were confined to isolated nuclei of differentiated cells, whereas high-grade lesions and invasive cancers showed elevated levels of equally distributed E7-specific signals in the cytoplasm of all tumour cells. The most abundant transcripts observed in intraepithelial neoplasias and in invasive cancers appear to initiate within ORF E7 and therefore have no coding potential for full-length E6 and E7 proteins. Our data show that the actual level of E7-specific transcripts in cancers is lower than anticipated from earlier studies using an ORF E6/E7-specific probe that hybridizes with the 5'-ends of the abundant mRNA class.

Base Sequence↗
PubMed1993

Expression of the different viral mRNAs of human papilloma virus 6 in a squamous-cell carcinoma of the bladder and the cervix.

The prevalence of HPV6 and HPV11 in benign condylomata or mild dysplasias has led to the view of HPV6/11 as rather harmless viruses in relation to carcinogenesis. However, the detection of HPV6/11 DNA in a number of individual cases of squamous-cell carcinomas of the anogenital/urinary tract could also point to a possible contribution of these viruses in the development of certain malignancies. Recently we have shown that the transcription of the E6 and E7 genes of HPV6 in benign anogenital condylomata is strictly confined to the basal cell layers of the epithelium, which express c-fos mRNA. This report describes the in situ hybridization analysis of individual mRNA species of HPV6 in 2 malignant tumours. A consistent feature of both carcinomas was the lack of detectable amounts of E6 mRNA, while the E7 mRNA was the major transcript observed. In situ hybridization with a riboprobe for c-fos revealed an expression pattern similar to that detected with the E7 probe. Hybridization with a probe specific for mRNA with a coding potential for a full-length E2 protein yielded weak signals in both carcinomas. Using restriction-enzyme analysis, we compared the long control region of HPV6 amplified by polymerase chain reaction from both tumours with already known HPV6 subtypes. In contrast to previous reports suggesting a correlation between genetic alterations in the long control region of HPV6 and increased malignant behaviour, our data do not support this hypothesis.

Base Sequence↗
PubMed1993

Human papillomavirus type 6 in squamous cell carcinoma of the bladder and cervix.

Human papillomavirus (HPV) types 16 and 18 are the most frequent genotypes identified in genital malignancies, while HPV types 6 and 11 are found predominantly in condylomas and low-grade dysplasias. It is thought that HPV types 16 and 18 represent high-risk genotypes, while HPV types 6 and 11 rarely, if ever, participate in the development of malignant tumors. In a series of over 300 invasive tumors of the lower genital tract analyzed for the presence of HPV three have been found to contain HPV type 6 DNA: two invasive squamous cell carcinomas of the cervix and one squamous cell carcinoma of the bladder. Human papillomavirus type 6 was the only HPV type detected in these tumor DNAs by Southern blot hybridization and by the polymerase chain reaction using both consensus and type-specific primers. In situ hybridization using whole genomic RNA probes localized viral DNA to tumor cells. Although extensive virologic and epidemiologic studies conducted in the last decade indicate that HPV types 16 and 18 are more likely to be associated with high-grade dysplasias and invasive cancer, HPV type 6 may not be as innocuous as previously supposed.

Adult↗
PubMed1992

Transcription of the E6 and E7 genes of human papillomavirus type 6 in anogenital condylomata is restricted to undifferentiated cell layers of the epithelium.

The E6 and E7 genes of human genital papillomaviruses (HPVs) appear to transform cells by different mechanisms. They seem to act synergistically but are not equally important when tested under diverse experimental conditions. We were therefore tempted to investigate the E6- and E7-specific transcription pattern in HPV6-infected condylomas separately, by in situ hybridization. Recent studies have identified three promoters within the E6-E7 region of HPV6 and HPV11 by applying S1, exonuclease VII, and cDNA analyses. On the basis of these data, we cloned subgenomic fragments of HPV6 into plasmid pBS to obtain riboprobes that differentiated between transcripts starting upstream of the E6 and E7 open reading frames, respectively. These different species of mRNAs were analyzed in serial thin sections of eight HPV6-positive anogenital condylomas. The E6 probe (nucleotides 7862 to 241) led to weak signals within the basal layer. In three cases, rather strong signals were confined to a few basal cells. The E7 probe (nucleotides 242 to 534) gave rise to a more pronounced labeling of all cells within the two to three lowest epidermal layers. In situ hybridization with a riboprobe for human c-fos revealed an expression pattern similar to that observed with the E7 probe. In contrast to the preferential expression of the transforming E6 and E7 genes in the lower epithelium, the major transcriptional activity of the virus was detected in the middle and upper third by probes colinear with the 3' moiety of the early region.

Adolescent↗
PubMed1990

The E7 protein of human papillomavirus 8 is a nonphosphorylated protein of 17 kDa and can be generated by two different mechanisms.

The E7 protein of human papillomavirus (HPV) 8 shows no in vitro transforming activity, in contrast to E7 of the genital HPVs, but seems to be involved in the control of viral DNA replication. To determine whether functional differences between the E7 proteins of HPV16 and HPV8 were reflected in differences in their biochemical properties, we characterized the E7 protein of HPV8 expressed from the late simian virus 40 promoter in COS 7 cells. An E7-specific antiserum was obtained by immunizing rabbits with a beta-gal-E7 fusion protein containing all of the E7 polypeptide. This antiserum specifically precipitated from [35S]cysteine but not from 32PO4-labeled transiently transfected COS 7 cells a protein with a low mobility of 17 kDa in SDS-PAGE. The high sedimentation rate in nondenaturing glycerol gradients pointed to an interaction with other proteins or to the existence of E7 oligomers. An association with the retinoblastoma protein could, however, be excluded. The 5' ends of HPV8 transcripts derived from the E6-E7 region in G418 selected rodent cells, which were mapped by nuclease S1 digestion, are located upstream of ORFs E6 and E7, respectively. No evidence for an E6*I-like mRNA was found. In COS 7 cells transfected with a plasmid expressing the E6-E7 region of HPV8 under the control of the late SV40 promoter, however, the E7 protein was translated from a polycistronic mRNA potentially encoding E6 and E7. These data indicate that the E7 protein of HPV8 may be expressed in two ways: (i) through translation of an E7-specific mRNA, as with HPV6 and HPV11, or (ii) through internal initiation of translation at the ATG of E7.

Animals↗
PubMed1988

Involvement of human papillomavirus type 8 genes E6 and E7 in transformation and replication.

We investigated the transforming activity of human papillomavirus type 8 (HPV8) by expressing all early open reading frames from a heterologous promoter in different rodent fibroblast lines. Morphological transformation was observed only in G418-selected mouse C127 and Rat 1 cells containing an intact E6-coding region. E6 of HPV8 did not transform NIH 3T3 cells as did E6 of bovine papillomavirus type 1. C127 cells transformed by E6 were anchorage independent and had a reduced serum requirement but did not form tumors in nude mice. E7 of HPV8 showed no transforming potential, in contrast to E7 of HPV18 and HPV16. It was, however, able to complement an E7 mutant of bovine papillomavirus type 1 with a defect in high-copy-number DNA maintenance. The data indicate that the expression of the HPV8 E6 open reading frame is sufficient to induce morphological transformation in rodent fibroblasts, whereas E7 is involved in the replication of the viral DNA.

Amino Acid Sequence↗
PubMed1987

Enhancement of Epstein-Barr virus membrane protein (LMP) expression by serum, TPA, or n-butyrate in latently infected Raji cells.

The BamHI Nhet region of the EBV DNA is known to code for two proteins. One is a membrane protein (LMP) with an apparent molecular weight of 60,000 on SDS-PAGE which is expressed in latently EBV infected cells. The second protein, so far unidentified, is presumably a late protein with a calculated molecular weight of 28,000 Da. Antisera against both proteins were generated by immunizing rabbits with either a fusion protein containing 155 amino acids of the C-terminus of LMP and a 37,000 mol wt piece of the bacterial anthranilate synthase or with a C-terminal synthetic peptide of 7 amino acids. These sera reacted with a protein varying in size between 60,000 and 65,000 mol wt on SDS-PAGE, found in all cell lines harboring EBV. In addition, these sera identified a second protein with an apparent molecular weight of 49,000 on SDS-PAGE in B95-8, P3HR-1, and M-ABA cells, which is presumably identical with the 28,000-Da protein mentioned above. Furthermore, with these sera a positive cytoplasmic immunofluorescence in 1 to 10% of the cells was obtained, depending on the cell line examined. Analyzing the nonproducer Raji cell line, the number of immunofluorescence-positive cells and the amount of the 60,000 protein, as judged by immunoblotting, was rapidly increased by addition of fresh medium with 10% fetal calf serum as well as by the tumor promoter TPA or to an even higher extend by n-butyrate. The kinetics of induction reached a maximum 24 hr after addition of medium plus 10% fresh serum or TPA or n-butyrate and decreased after 24 to 48 hr. Since the induction of the EBV early antigen (EA) associated proteins by TPA or n-butyrate exhibits a diverse kinetic with a maximum at 72 hr, the regulation of the 60,000 protein synthesis appears to be different from known EA-associated proteins.

Antigens, Viral↗
PubMed1987

Genetic relationship among human papillomaviruses associated with benign and malignant tumours of patients with epidermodysplasia verruciformis.

Human papillomaviruses (HPVs) 5, 8, 19 and 25 induce macular skin lesions in patients with epidermodysplasia verruciformis. HPVs 5 and 8 are known to prevail in skin carcinomas, which develop in about one-third of these patients. We compared the viral DNAs by heteroduplex analysis and on the basis of partial nucleotide sequences. The colinear genomes were closely related and showed nucleotide sequence homology in the range of 40% to 90%. Homology values between 70% and 80% were observed throughout roughly 4 kb of the heteroduplex molecules as estimated after 'calibration' by partial sequencing. Heteroduplex analysis did not support the hypothesis that the different HPV types arise by recombination and allowed no grouping to correlate with the association with malignant tumours. The upstream regulatory sequences of HPVs 8, 19 and 25 appeared highly conserved but differed from those of previously sequenced papillomaviruses in length, in the TATA motif, in the copy numbers and positions of the ACCGN4CGGT palindrome and of a YGCCAA direct repeat, and in two strictly conserved blocks of 33 and 29 nucleotides.

Base Sequence↗
PubMed1986

Classification of the papillomaviruses--mapping the genome.

Papillomaviruses form one genus of the Papovaviridae family. They share common antigenic determinants and their DNAs cross-hybridize under conditions of low stringency. The classification of papillomaviruses is at present based on the host range and the relatedness of the nucleic acids. Isolates are considered independent types if there is less than 50% cross-hybridization in the liquid phase according to a standard protocol. At least 31 human and six bovine papillomavirus types can be differentiated on this basis. The host range does not reflect the natural relationship between the viruses. Subgenera, which differ in biological properties, can be distinguished in outline. Data on overall sequence homology are insufficient for a meaningful classification because two types of virus may be closely related within one genome region and rather heterogeneous in other areas. Some new isolates appear as intermediates between previously well-separated types and complicate the system. A reasonable classification of such types of papillomavirus should be based on homologies between genes that are relevant for differences in the biology of the viruses. A functional mapping of the rather uniformly organized, colinear genomes of papillomaviruses has been started. Genetic studies with bovine papillomavirus type 1 have assigned functions in replication, transformation, gene expression and capsid synthesis to individual open reading frames.

Amino Acid Sequence↗
PubMed1986

Epidermodysplasia verruciformis-associated human papillomavirus 8: genomic sequence and comparative analysis.

Human papillomavirus (HPV) 8 induces skin tumors which are at high risk for malignant conversion. The nucleotide sequence of HPV8 has been determined and compared to sequences of papillomaviruses with different oncogenic potential. The general organization of the HPV8 genome is similar to that of other types. Highly conserved, genus-specific sequences were found in open reading frames (ORFs) E1, E2, and L1. In ORFs E6, E7, and L2, HPV8 is more distantly related, but it was possible to differentiate subgenera in which HPV8 belonged to the HPV1-cottontail rabbit papillomavirus group. Sequences within ORF E4 and part of ORF L2 are rather type specific. HPV8 stands out by several unique features: the considerably reduced size of the noncoding region (397 base pairs), with a seemingly low potential for forming complex secondary structures; a cluster of putative promoter elements in the 3' half of ORF E1; an RNA polymerase III promoter-like sequence close to the C terminus of ORF E2; and of particular interest, the homology between the putative protein encoded by ORF E4 and the Epstein-Barr virus nuclear antigen 2 protein, which may reflect similar mechanisms in virus-mediated transformation.

Amino Acid Sequence↗