Types 2c Research Articles

Analysis of human papillomavirus types in exophytic condylomata acuminata by hybrid capture and Southern blot techniques.

Exophytic condylomata acuminata of the external genitalia of 40 patients were analyzed for human papillomavirus (HPV) DNA by the Southern blot and hybrid capture methods. All lesions were initially analyzed by the Southern blot method by using a mixture of HPV type 6, 11, 16, and 18 whole genomic probes. Southern blots demonstrated characteristic PstI restriction patterns of HPV type 6, 11, or 16 in all but one lesion. HPV 6 subtypes accounted for 28 of 39 HPV-positive lesions. Twenty-seven of these 28 lesions contained HPV type 6a, and 1 lesion contained HPV type 6c. Eight lesions contained HPV type 11 and three contained HPV type 16. Two of the three condylomata acuminata containing HPV type 16 were obtained from solid-organ transplant recipients receiving immunosuppressive medications. The third lesion containing HPV type 16 was a typical exophytic condyloma acuminatum from a woman with previously resected vulvar carcinoma. The hybrid capture assay detected HPV DNAs in all lesions except the Southern blot-negative lesion. Twenty-five lesions were positive for the A probe only (HPV types 6 and 11 and related types). All of these lesions were found to contain HPV type 6 or 11 sequences in the Southern blot assay. The remaining 14 lesions were positive for both the A probe and the B probe (HPV types 16 and 18 and related types). The strongest signal in these 14 lesions by the hybrid capture assay was consistent with the result of the Southern blot assay in all but one case. We conclude that (i) HPV type 6a is the most common type found in these lesions, (ii) HPV type 16 may be present more often in exophytic condylomata acuminata from immunosuppressed individuals, (iii) hybrid capture is a useful tool for documenting the presence of HPV sequences in DNAs from exophytic condylomata acuminata, and (iv) in samples containing multiple HPV types, hybrid capture allows detection of minority HPV types.

Detection by antibody probes of human papillomavirus type 6 E5 proteins in respiratory papillomata

We have demonstrated the expression of proteins arising from the E5a and E5b open reading frames (ORFs) of human papillomavirus type 6c (HPV-6c) in respiratory tract papillomata. Recombinant plasmids were constructed to express the ORFs in the bacterial vectors pATH and pRIT2T. Fusion proteins were purified and injected into rabbits to produce polyclonal antibodies. Characterized antibodies generated against these fusion proteins were used in immunoperoxidase assays to identify the presence and distribution of HPV-6 E5 proteins in biopsy specimens of respiratory tract papillomata. The results showed that the E5a and E5b proteins were distributed throughout the thickness of the epithelium in the papillomata but not in the basal layer. The proteins were found in nuclei and in the cytoplasm of koilocytotic cells. Positive reactivity with a similar distribution in the epithelium and subcellular location was obtained in papillomata induced by other HPV-6 subtypes. This cross-reactivity was not unexpected, since nucleotide and amino acid (aa) sequence comparisons between HPV-6c and -6e demonstrated 79% sequence identity with 15 aa substitutions in the 91 aa of E5a. The E5b ORF of HPV-6c has the potential to encode a protein of 74 aa that differed at 28 positions compared with the 72 aa of HPV-6e.

Structural analysis of human papillomavirus type 6c isolates from condyloma acuminatum and juvenile-onset and adult-onset laryngeal papillomata

The human papillomavirus type 6c (HPV-6c) genome was molecularly cloned from biopsy specimens of a juvenile-onset and an adult-onset respiratory-tract papillomata and a condyloma acuminatum of the cervix. To determine if the genital-tract isolate and respiratory-tract isolates contain divergent sequences that may account for a difference in tissue trophism or for a difference in the age of onset of the disease, fine-structure mapping, heteroduplex analysis by electron microscopy, and nucleotide sequencing were used to examine the sequence relationship among these HPV-6c isolates. No differences were found in the digestion patterns with 23 restriction enzymes. Heteroduplex analysis among the three genomes demonstrated that they were colinear without apparent deletions or rearrangements and had greater than 90% sequence identity. In heteroduplex analyses with a different subtype (HPV-6e) that was molecularly cloned from a genital wart, the genomes were colinear with greater than 90% sequence identity over 90% of their length. The most divergent region had 75-80% sequence identity and was localized to the part of the genome containing the E5a and E5b open reading frames (ORFs). Comparison of the sequence of 1430 nucleotides in this region for two of the HPV-6c isolates did not identify any differences between them. Comparison with the published sequences of HPV-6b identified deletions/insertions and base changes with approximately 75% sequence identity, and comparison with HPV-11 identified only six base changes. Conservation of sequences in the E4-E5 region and similarity in the restriction enzyme maps demonstrated that HPV-6c and HPV-11 are independent isolates of the same HPV-6 subtype.

Morphological evidence for a probable secretory site of the male sex pheromones of Nauphoeta cinerea (blattaria, blaberidae). 2. electron microscope studies.

Light and electron microscopy of the glandular epithelium of intersegmental membranes between sternites three and seven and tergites two and eight of various age groups of Nauphoeta cinerea male adults and one age group of female adults discloses differences in the epithelia of the intersternite and intertergite. The intersternal epithelium appears thicker, more glandular, and stratified. Altogether, seven cell types are recognizable, six in the male and two in the female. They are designated as types 1, 2a, 2b, 2c, 3, 4, and 5. Of these, types 1, 2a, 3, and 4 are recognizable on the sternum; types 1, 2b, and 5 on the tergum of the mature male integuments. Types 1 and 2c are found on the sternum of mature female. The cell types undergo morphological differentiation after adult emergence and show different stages of secretory activity. Type 1 are squamous cuticle-secreting cells; type 2a, 2b, and 2c are columnar-glandular and contain electron-transparent secretory vesicles of various sizes, which increase greatly in number and size in the 5-day-old adult males when the glands are most active. The vesicular size and number also differ between types 2a, 2b, and 2c cells of the same age group. The vesicles are assumed to be derived from smooth endoplasmic reticulum. The type 2 gland cells are also provided with a secretory end apparatus lined by cuticle and bordered by microvilli through which the secretion is believed to be released by exocytosis. The end apparatus leads into a cuticular ductule that opens to the surface of the cuticle as a cup-shaped receptacle, which is more conspicuous in the male intersternite. In the active gland cells, the mitochondria near the end apparatus are swollen and vacuolated. Type 3 cells are seen only on the intersternum and are believed to secrete the cuticular ductule that proceeds from the end apparatus. Type 4 cells are also recognizable only on the male intersternum and contain closely packed, electron-dense bodies, which are most numerous in mature (5-day-old) males. Type 5 cells with their dense cytoplasm are located basally in the intertergal epithelium. The functional significance of type 4 and 5 cells in the males and type 2c cells in the female is not clear. On the basis of differences in morphology, pheromone activity, and sexual behavior, it is suggested that the pheromones secreted by the intersternal and intertergal glands in the male are different, the former secreting a seducin that attracts the female to the male and the latter an "aphrodisiac" acting as a contact pheromone important in accomplishing mating.

Immunological characterization of phosphoprotein phosphatases

Phosphoprotein phosphatases regulate the biological activities of proteins through their involvement in cyclic phosphorylation/dephosphorylation cascades. A variety of multimeric phosphatases have been isolated and grouped into several classes, termed type 1 and types 2A, 2B, and 2C. To elucidate the relationship between the different phosphoprotein phosphatases, highly purified enzymes from soil amoebae, turkey gizzards, bovine heart and brain, and rabbit skeletal muscle and reticulocytes were tested for immunological antigenic relatedness. Two heterologous antibody preparations were employed for this purpose. One was made against an Acanthamoeba type 2A phosphatase and the other was made to bovine brain phosphatase type 2B (calcineurin, holoenzyme). Specific subunit cross-reactivity was examined by protein blot (“Western”) analysis. The antibody to the type 2A phosphatase reacted with the catalytic subunits of every type 2 enzyme tested, including both the catalytic and Ca 2+-binding subunits of the Ca 2+/calmodulin-dependent type 2B phosphatase (calcineurin), bovine cardiac type 2A phosphatase, and turkey gizzard smooth muscle phosphatase-1 (type 2A 1). It did not react with any type 1 phosphatase (catalytic subunit or ATP-Mg-dependent). The antigenic relatedness of calcineurin and the bovine cardiac type 2A phosphatase ( M r 38,000) was demonstrated further by protein blot analysis showing that the anti-calcineurin antibody cross-reacted with both enzymes. The mutual cross-reactivity poses an intriguing problem because these enzymes are so different in their molecular structures and modes of regulation. The degree of evolutionary conservation exhibited by the antigenic cross-reactivity of the type 2 enzymes from a broad range of species and tissues suggests a strong selective pressure on maintaining one or more features of these important regulatory enzymes.

Isolation and characterization of six new genome types of human adenovirus types 1 and 2

Several of the 41 types of human adenovirus have been divided into genome types based on aberrant restriction endonuclease digestion patterns of viral DNA. In the process of screening a large number of clinical adenovirus isolates by restriction endonuclease digestion of viral DNA, we have identified nine isolates from eight patients which were identified by neutralization as adenovirus type 1 or 2 but had aberrant cleavage patterns. Cleavage sites for the enzymes SmaI, EcoRI, HindIII, KpnI, and HpaI were mapped. The variants could be placed in six distinct groups based on cleavage patterns. The designation genome types 1a, 1b, 1c, 2b, 2c, and 2d are proposed for these isolates.

Iodine-concentrating cells in the endostyle of Ammocoetes.

The iodine-concentrating cells in the endostyle were examined in four larvae of petromyzon planeri. The visceral cells (types 2c, 3, and 4) were provided with cilia and contained granules with a characteristic content of concentric lamellae. Type 3 was particularly rich in ergastoplasm. The parietal cells (type 5) were short with a polymorphous content indicating a resorptive rather than a synthetic function. Based on the ultrastructure of the cells it has not been possible to decide which group forms the follicular epithelium in the petromyzone after metamorphosis, but it is most likely type 3 with its protein-synthetizing apparatus.