Two Polypyrimidine Tracts in Intron 4 of the Major Immediate Early Gene Are Critical for Gene Expression Switching from IE1 to IE2 and for Replication of Human Cytomegalovirus.

UNLABELLED: The human cytomegalovirus (HCMV) major immediate early (MIE) gene is essential for viral replication. The most abundant products encoded by the MIE gene include IE1 and IE2. Genes of IE1 and IE2 share the MIE promoter (MIEP), the first 3 exons, and the first 2 introns. IE1 is expressed earlier than IE2 after CMV infection or MIE gene transfection. In this study, we identified 2 polypyrimidine (Py) tracts in intron 4 (between exons 4 and 5) that are responsible for transcriptional switching from IE1 to IE2. The first Py is important and the second one is essential for the splicing and expression of IE2. In searching for the mechanisms of MIE gene switching from IE1 to IE2, we found that the second Py was required for the IE2's fourth intron to bind to a splicing factor such as U2AF65, as determined by an RNA electrophoretic mobility shift assay and a chromatin immunoprecipitation (ChIP) assay, while the first Py enhanced the binding of U2AF65 with the intron. An HCMV BACmid with the second Py mutated failed to produce any virus, while the HCMV with the first Py mutated replicated with a defective phenotype. Furthermore, we designed a small RNA (scRNAPy) that is complementary to the intron RNA covering the two Pys. The scRNAPy interfered with the interaction of U2AF65 with the intron and repressed the IE2 expression. Therefore, our studies implied that IE2 gene splicing might be an anti-CMV target. IMPORTANCE: CMV is a ubiquitous herpesvirus and a significant cause of disease and death in the immunocompromised and elderly. Insights into its gene regulation will provide clues in designing anti-CMV strategies. The MIE gene is one of the earliest genes of CMV and is essential for CMV replication. It is known that the MIE gene needs to be spliced to produce more than two proteins; however, how MIE gene splicing is regulated remains elusive. In the present studies, we identified two Pys in intron 4 and found that the first Py is important and the second is required for the splicing and expression of IE2. We further investigated the mechanisms of gene switching from IE1 to IE2 and found that the two Pys are responsible for U2AF65's binding with intron 4. Therefore, the Pys in intron 4 are the cis elements that determine the fate of IE2 splicing. Furthermore, we found that a small RNA that is complementary to intron 4 repressed IE2 expression. Hence, we provide the first piece of evidence for a unique mechanism of MIE gene regulation at the splicing level.

Bilateral Spigelian Hernias Robotic Repair: A Novel Approach to a Rare Surgical Entity.

Spigelian hernias are an uncommon protrusion defect noted between the rectus abdominis and the transversus abdominis muscles, at the semilunar line, with a low incidence of approximately 0.12% to 2% of all ventral hernias. Furthermore, the incidence of bilateral cases is noted to be even lower in the general population. They are associated with bowel incarceration and strangulation; hence surgical repair is indicated. Therapeutic alternatives for Spigelian hernias include open repair, however laparoscopic repair stands as the standard of care in these cases. Feasible and successful robotic repair has been reported, with associated advantages in terms of visualization and surgical instrument dexterity. We present the first ever reported cases of bilateral Spigelian hernias repaired using robotic approach. Two female Puerto Rican patients referred to our institution complaining of abdominal pain, where imaging studies found bilateral Spigelian hernias. In both cases, a transabdominal preperitoneal repair was performed using the Da Vinci Surgical System. Both patients were discharged home tolerating oral intake with adequate wound healing. On follow up visits, patients denied abdominal discomfort and had adequate wound healing. Robotic surgery for Spigelian hernias poses an advantage over laparoscopic repair as improved visualization, mobility, and precision in movements allow for more gentle tissue manipulation. Furthermore, this is the first evidence of safe and effective repair in the uncommon entity of bilateral cases, providing a newer alternative in the setting of such presentation.

[Knowledge and attitude of adolescent girls regarding folic acid use]. / Conocimiento y actitud de las adolescentes en el consumo del ácido fólico.

OBJECTIVE: To determine the adolescent's knowledge and attitude on the use of folic acid to avoid congenital neural tube defects. METHOD: Three hundred and three (303) adolescent students from two different schools, one private and one public, were asked to complete a pre-test survey. After completing the survey, a lecture was given addressing the importance of folic acid and the benefits of taking this vitamin at an early stage to avoid neural tube defect. Two weeks later a post-test was administered to evaluate the knowledge acquired. The variables were grouped based on the knowledge of folic acid used, the attitudes toward folic acid consumption and the economic level of the participants. RESULTS: On the pre-test the adolescents demonstrated 52.5% of knowledge on the use of folic acid. After the educational conference, the tested knowledge increased to 87.5%. Eighteen percent (18%) of the adolescents reported on the pre-test that they consumed folic acid; this result increased to 69.3% on the post-test. On the pre-test, 31.7% of the participants related folic acid as an etiology of neural tube defect. Following the post-test, 96% of the adolescents were able to provide correct answers about the benefits of folic acid. CONCLUSION: Less than 50% of the adolescents recognized that folic acid is a vitamin. More than 80% of the surveyed adolescents do not consume folic acid. In general, the majority of the adolescents did not know the benefits of folic acid during the pre-conception period.