Identification of three functions of the adenovirus e4orf6 protein that mediate p53 degradation by the E4orf6-E1B55K complex.

Complexes containing adenovirus E4orf6 and E1B55K proteins play critical roles in productive infection. Both proteins interact directly with the cellular tumor suppressor p53, and in combination they promote its rapid degradation. To examine the mechanism of this process, degradation of exogenously expressed p53 was analyzed in p53-null human cells infected with adenovirus vectors encoding E4orf6 and/or E1B55K. Coexpression of E4orf6 and E1B55K greatly reduced both the level and the half-life of wild-type p53. No effect was observed with the p53-related p73 proteins, which did not appear to interact with E4orf6 or E1B55K. Mutant forms of p53 were not degraded if they could not efficiently bind E1B55K, suggesting that direct interaction between p53 and E1B55K may be required. Degradation of p53 was independent of both MDM2 and p19ARF, regulators of p53 stability in mammalian cells, but required an extended region of E4orf6 from residues 44 to 274, which appeared to possess three separate biological functions. First, residues 39 to 107 were necessary to interact with E1B55K. Second, an overlapping region from about residues 44 to 218 corresponded to the ability of E4orf6 to form complexes with cellular proteins of 19 and 14 kDa. Third, the nuclear retention signal/amphipathic arginine-rich alpha-helical region from residues 239 to 253 was required. Interestingly, neither the E4orf6 nuclear localization signal nor the nuclear export signal was essential. These results suggested that if nuclear-cytoplasmic shuttling is involved in this process, it must involve another export signal. Degradation was significantly blocked by the 26S proteasome inhibitor MG132, but unlike the HPV E6 protein, E4orf6 and E1B55K were unable to induce p53 degradation in vitro in reticulocyte lysates. Thus, this study implies that the E4orf6-E1B55K complex may direct p53 for degradation by a novel mechanism.

Identification and elimination of an aberrant splice product from cDNAs encoding the human adenovirus type 5 E4orf6 protein.

Growing awareness of the central role of the E4orf6 protein in regulating the infectious cycle of human adenoviruses has led to greatly intensified efforts to define its functions and mechanisms of action. Many workers employ cDNAs to generate plasmid or viral vectors to express E4orf6 in the absence of other viral products. In addition to the normal 34-kDa product, such vectors consistently produce a polypeptide of about 8 kDa. In the present report we show that this protein is produced by an aberrant mRNA utilizing the 5' splice donor site used normally by the virus to produce the E4orf6/7 product, which shares 58 residues with E4orf6. This amino terminal coding sequence is linked to a 3' sequence via a novel splice acceptor site in an alternative reading frame of the E4orf6 cDNA. The 5' donor site was altered by PCR-directed mutagenesis to yield a construct that produces high levels of E4orf6 in the absence of the 8-kDa polypeptide. This construct should eliminate some of the problems encountered previously using the wild-type E4orf6 coding sequence.

An unusual cause of cervical lymphadenitis: Kikuchi-Fujimoto disease.

Cervical lymphadenitis may be the result of diverse conditions in a patient. Clinical and epidemiologic information about cervical lymphadenitis can often lead to a presumptive diagnosis and, thus, limit the number of studies required as well as direct the type of initial therapy administered. We report a case of cervical lymphadenitis in a Vietnamese woman for whom a presumptive diagnosis of tuberculosis was made and antituberculous therapy was started. Pathologic examination of an excised lymph node revealed the correct diagnosis--histiocytic necrotizing lymphadenitis, or Kikuchi-Fujimoto disease. We review the clinical, epidemiologic, and pathologic features of this recently recognized disease. Careful examination of excised material from the lymph nodes should prevent patients who have Kikuchi-Fujimoto disease from receiving unnecessary treatment.

Foreign bodies in the tracheobronchial tree--a retrospective study of 217 cases.

This is a retrospective study of 217 cases of foreign body in the tracheobronchial tree. The diagnostic techniques, current management, and statistics related to sex, age, yearly incidence, duration of symptoms, nature of foreign body, location, and duration of hospitalization are discussed. The most common complications resulting from a foreign body in the tracheobronchial tree were atelectasis, subglottic stenosis, and pneumonia.