PubMed Health⌕ Search

Biomedical subjects

Mutsuhito Ohno

Publications and source records attributed to Mutsuhito Ohno.

9 recordsLinked to original sources
PubMed2006

Binding of a novel SMG-1-Upf1-eRF1-eRF3 complex (SURF) to the exon junction complex triggers Upf1 phosphorylation and nonsense-mediated mRNA decay.

Nonsense-mediated mRNA decay (NMD) is a surveillance mechanism that degrades mRNA containing premature termination codons (PTCs). In mammalian cells, recognition of PTCs requires translation and depends on the presence on the mRNA with the splicing-dependent exon junction complex (EJC). While it is known that a key event in the triggering of NMD is phosphorylation of the trans-acting factor, Upf1, by SMG-1, the relationship between Upf1 phosphorylation and PTC recognition remains undetermined. Here we show that SMG-1 binds to the mRNA-associated components of the EJC, Upf2, Upf3b, eIF4A3, Magoh, and Y14. Further, we describe a novel complex that contains the NMD factors SMG-1 and Upf1, and the translation termination release factors eRF1 and eRF3 (SURF). Importantly, an association between SURF and the EJC is required for SMG-1-mediated Upf1 phosphorylation and NMD. Thus, the SMG-1-mediated phosphorylation of Upf1 occurs on the association of SURF with EJC, which provides the link between the EJC and recognition of PTCs and triggers NMD.

Codon, Nonsense↗
PubMed2005

Shigella effector IpaH9.8 binds to a splicing factor U2AF(35) to modulate host immune responses.

Shigella effectors injected into the host cell via the type III secretion system are involved in various aspects of infection. Here, we show that one of the effectors, IpaH9.8, plays a role in modulating inflammatory responses to Shigella infection. In murine lung infection model, DeltaipaH9.8 mutant caused more severe inflammatory responses with increased pro-inflammatory cytokine production levels than did wild-type Shigella, which resulted in a 30-fold decrease in bacterial colonization. Binding assays revealed that IpaH9.8 has a specific affinity to U2AF(35), a mammalian splicing factor, which interferes with U2AF(35)-dependent splicing as assayed for IgM pre-mRNA. Reducing the U2AF(35) level in HeLa cells and infecting HeLa cells with wild-type caused a decrease in the expression of the il-8, RANTES, GM-CSF, and il-1beta genes as examined by RT-PCR. The results indicate that IpaH9.8 plays a role in Shigella infection to optimize the host inflammatory responses, thus facilitating bacterial colonization within the host epithelial cells.

Animals↗
PubMed2005

Mutations conferring resistance to human immunodeficiency virus type 1 fusion inhibitors are restricted by gp41 and Rev-responsive element functions.

One of the human immunodeficiency virus (HIV) envelope proteins, gp41, plays a key role in HIV fusion. A gp41-derived peptide, T-20, efficiently inhibits HIV fusion and is currently approved for treatment of HIV-infected individuals. Although resistant variants have been reported, the mechanism of the resistance remains to be defined. To elucidate the mechanism in detail, we generated variants resistant to C34, a peptide derived from the gp41 carboxyl terminus heptad repeat (C-HR) in vitro. The resistant variants had a 5-amino-acid deletion in gp120 and a total of seven amino acid substitutions in gp41. Binding assays revealed that an I37K substitution in the N-terminal heptad repeat (N-HR) impaired the binding of C34, whereas an N126K substitution in the C-HR enhanced the binding to mutated N-HR, indicating that both mutations were directly involved in resistance. On the other hand, substitutions for A30 and D36 seemed to be secondary mutations, located complementary to each other in the Rev-responsive element (RRE), and were mutated simultaneously to maintain the secondary structure of the RRE that was impaired by the mutations at I37. Thus, HIV acquired resistance to C34 by mutations in N-HR, which directly interacted with C34. However, since this region also encoded the RRE, additional mutations were required to maintain viral replication. These results suggest that HIV fusion is one of the attractive targets for HIV chemotherapy.

Amino Acid Sequence↗
PubMed2004

RNA length defines RNA export pathway.

Different RNA species are exported from the nucleus by distinct mechanisms. Among the different RNAs, mRNAs and major spliceosomal U snRNAs share several structural similarities, yet they are exported by distinct factors. We previously showed that U1 snRNAs behaved like an mRNA in nuclear export if various approximately 300-nucleotide fragments were inserted in a central position. Here we show that this export switch is dependent on the length of the insertion but independent of its position, indicating unequivocally that this switch is indeed the result of RNA length. We also show that intronless mRNAs can be progressively converted to use the U snRNA export pathway if the mRNAs are progressively shortened by deletion. In addition, immunoprecipitation experiments show that the protein composition of export RNPs is influenced by RNA length. These findings indicate that RNA length is one of the key determinants of the choice of RNA export pathway. Based on these results and previous observations, a unified model of how an RNA is committed to a specific export pathway is proposed.

Active Transport, Cell Nucleus↗
PubMed2004

SR proteins preferentially associate with mRNAs in the nucleus and facilitate their export to the cytoplasm.

Different classes of RNA are exported to the cytoplasm by distinct mechanisms. Each class of RNA forms distinct complexes with nuclear proteins prior to its export to the cytoplasm. In our attempt to obtain comprehensive information of protein factors that specifically associate with mRNAs in the nucleus, we performed in vivo UV-crosslinking analysis after microinjection of various RNAs into Xenopus oocyte nucleus. We found a group of proteins preferentially crosslinked to mRNAs. Immunoprecipitation experiments revealed that some of the crosslinked signals corresponded to SR (serine/arginine-rich) proteins, a family of essential RNA-binding proteins involved in pre-mRNA splicing. It was previously suggested that some members of SR protein family are involved in export of a specific intronless mRNA, histone H2A mRNA and some spliced mRNAs. However, it is still to be clarified if SR proteins are involved in export of general mRNAs, especially general intronless mRNAs that do not contain specific RNA export elements. When we microinjected an antibody against SR proteins into the nucleus, export of mRNAs was severely inhibited, regardless of whether the mRNAs were produced via pre-mRNA splicing or not, whereas export of other RNAs was not affected. These results unequivocally showed that SR proteins are involved in export of both general intronless and spliced mRNAs.

Animals↗
PubMed2002

Identity elements used in export of mRNAs.

Different classes of RNA are exported from the nucleus by distinct factors. We demonstrate that U1 snRNA is exported like an mRNA on insertion of a pre-mRNA intron or either sense or antisense mRNA exon sequences. mRNA-specific factors are recruited onto the spliced or elongated U1 RNA whereas U snRNA-specific factors are not, suggesting that an unstructured region of sufficient length in an RNA acts as a dominant determinant of mRNA identity. After export, spliced U1 RNA undergoes cytoplasmic maturation but is not reimported into the nucleus. These data provide insight into mechanisms for discrimination of different classes of nuclear RNA and demonstrate that two RNAs of identical sequence can have distinct cytoplasmic fates depending on their mode of export.

Active Transport, Cell Nucleus↗