No abstract

An internal ribosome entry site (IRES) has been used to facilitate the expression of more than one protein in a single transcript. In this study, we examined the translational activities of several IRES elements derived from different RNA viruses. The protein expression of encephalomyocarditis virus (EMCV) IRES is similar to that of hepatitis C virus (HCV) IRES in mammalian cells. Notably, the protein expression of enterovirus 71 (EV71) IRES was 23-fold higher than the efficiency of EMCV IRES following normalization of mRNA transcriptional level. Thus, expression of the secreted alkaline phosphatase (SEAP) reporter protein in mammalian cells may be controlled at desirable levels by using appropriate IRES in the expression vector.

The HCMV IE2 protein negatively autoregulates its own expression as well as represses the transactivation activity of p53. Using the repression domain of IE2 as bait in the yeast two-hybrid system, Nrf1 and Nrf2, members of the CNC-bZIP family, were found to be IE2-interacting proteins. Residues 331-448 encompassing the DNA-binding and the dimerization domains of Nrf1 are sufficient for the interaction. The interaction was further confirmed in vitro by a glutathione S-transferase pull-down assay and in vivo by co-immunoprecipitation. In transient transfection studies, transcription driven by six copies of an NF-E2 site or by chimeric proteins between the DNA-binding domain of LexA and members of the CNC-bZIP family is repressed by IE2. Importantly, the DNA binding activity of the Nrf1/MafK heterodimer is not impeded by IE2. In a parallel study, CNC-bZIP factors attenuate the negative autoregulation of IE2. The attenuation could be explained by the finding that Nrf1 functions alone and synergistically with its heterodimerization partner, MafK, in inhibiting the DNA binding activity of IE2. Taken together, these results demonstrate the existence of antagonism between members of the CNC-bZIP family and IE2.Human cytomegalovirus (HCMV), 1 a member of the ␤ subgroup of herpesviruses, has a double-stranded DNA genome of 229,354 base pairs with a potential to encode more than 200 proteins (1). A number of immediate-early (IE) proteins of HCMV are made following entry of the virus into cells (2). Among them the IE2 86-kDa protein (referred hereafter as IE2) is most studied. IE2 appears to be a promiscuous transactivator of viral and cellular gene expression (3-5). IE2 stimulates transcription by interacting with general transcription factors and/or gene-specific factors (6 -9). In addition, IE2 negatively autoregulates its own expression by binding to a short nucleotide sequence, termed the cis repression signal (CRS), located immediately downstream of the TATA box of the HCMV major immediate early promoter (MIEP) (10, 11). Binding of IE2 to CRS prevents the recruitment of RNA polymerase II to the promoter via steric hindrance (12). Thus, IE2-mediated, CRSdependent transcriptional repression is both passive and position-dependent. The efficiency of IE2-mediated repression of MIEP seems to be cell-dependent (4, 10, 13), strongly suggesting that the IE2 activity may be modulated by post-translational modification and/or by interaction with cellular proteins. Indeed, phosphorylation has been shown to be one of these factors (14).Furthermore, recent studies demonstrate that IE2 can be converted into a trans-repressor following binding to p53 (15,16). A transcriptional repression domain has been mapped to the C terminus of IE2 (16), which is also required for its transcriptional activation, DNA binding, and interaction with CREB, c-Jun, RB, TBP, and TFIIB (6 -9). The DNA-binding and the repression domains of IE2 seem to be overlapping, because an IE2 mutant devoid of DNA binding activity also loses its repression activity (1...

Ehlers-Danlos syndrome (EDS) includes a group of connective tissue disorders with abnormal collagen metabolism and a diverse clinical spectrum. We report two siblings with EDS who both presented with congenital diaphragmatic hernia (CDH). The elder sister suffered from recurrent diaphragmatic hernia twice and EDS was overlooked initially. Echocardiography as well as contrast-enhanced magnetic resonance angiography (MRA) showed dilatation of the pulmonary artery, and marked elongation and tortuosity of the aorta and its branches. A diagnosis of EDS was eventually established when these findings were coupled with the clinical features of hyperelastic skin. Her younger brother also had similar features. This report emphasizes that EDS may present as CDH in a small child which could easily be overlooked. Without appropriate surgery, diaphragmatic hernia might occur. Echocardiographic screening is recommended in patients with CDH. Contrast-enhanced MRA can be helpful in delineation of abnormally tortuous aortic great vessels that are an important clue to the early diagnosis of EDS.