Biomedical subjects

Ming-Chih Lai

Publications and source records attributed to Ming-Chih Lai.

6 recordsLinked to original sources

PubMed2004

Hypophosphorylated ASF/SF2 binds TAP and is present in messenger ribonucleoproteins.

Serine/arginine-rich proteins (SR proteins) function in precursor mRNA (pre-mRNA) splicing and may also act as adaptors for mRNA export. SR proteins are dynamically phosphorylated in their RS domain, and differential phosphorylation modulates their splicing activity and subcellular localization. In this study, we investigated the influence of phosphorylation on the function of SR proteins in events occurring during mRNA maturation. Immunoprecipitation experiments showed that the mRNA export receptor TAP associates preferentially with the hypophosphorylated form of shuttling SR proteins, including ASF/SF2. Overexpression of ASF induced subnuclear relocalization of TAP to SR protein-enriched nuclear speckles, suggesting their interaction in vivo. Moreover, the ASF found in a nucleoplasmic fraction rich in heterogeneous nuclear ribonucleoprotein (hnRNP) complexes is hyperphosphorylated, whereas mature messenger RNP (mRNP)-bound ASF is hypophosphorylated. Therefore, hypophosphorylation of ASF in mRNPs coincides with its higher affinity for TAP, suggesting that dephosphorylation of ASF promotes both its incorporation into mRNPs and recruitment of TAP for mRNA export. Thus, the phosphorylation state of RS domains may modulate the function of mammalian shuttling SR proteins during mRNA maturation or export.

Base Sequence↗

PubMed2003

Efficient algorithm on a nonstaggered mesh for simulating Rayleigh-Bénard convection in a box.

An efficient semi-implicit second-order-accurate finite-difference method is described for studying incompressible Rayleigh-Bénard convection in a box, with sidewalls that are periodic, thermally insulated, or thermally conducting. Operator-splitting and a projection method reduce the algorithm at each time step to the solution of four Helmholtz equations and one Poisson equation, and these are solved by fast direct methods. The method is numerically stable even though all field values are placed on a single nonstaggered mesh commensurate with the boundaries. The efficiency and accuracy of the method are characterized for several representative convection problems.

Journal Article↗

PubMed2003

Fatty acids modulate transforming growth factor-beta activity and plasma clearance.

The activity and plasma clearance of transforming growth factor (TGF)-beta are known to be regulated by activated alpha2-macroglobulin (alpha2M*). This has been implicated in pathophysiological processes, but no small molecule compounds have been reported to modulate TGF-beta activity by affecting the interaction of TGF-beta and alpha2M*. Here, we demonstrate that fatty acids are capable of inhibiting complex formation of TGF-beta isoforms and alpha2M* as demonstrated by nondenaturing and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. This is dependent on carbon chain length (C20, C18, C16, C14 > C12 > C10), degree of unsaturation (polyunsaturated > saturated), and TGF-beta isoforms (TGF-beta1 > TGF-beta2 > TGF-beta3). Arachidonic acid, which is one of the most potent inhibitors, is also capable of dissociating TGF-beta-alpha2M* complexes, but higher concentrations are required. Arachidonic acid appears to inhibit TGF-beta-alpha2M* complex formation by binding specifically to alpha2M* as demonstrated by gel filtration chromatography. Arachidonic acid reverses the inhibitory effect of alpha2M* on TGF-beta binding, TGF-beta-induced growth inhibition, and TGF-beta-induced transcriptional activation in mink lung epithelial cells and affects plasma clearance of TGF-beta-alpha2M* complexes in mice. These results show that fatty acids are effective modulators of TGF-beta activity and plasma clearance and may be useful in treating human diseases through their effects on the interaction of TGF-beta and alpha2M*.

Animals↗

PubMed2003

Differential effects of hyperphosphorylation on splicing factor SRp55.

Members of the serine/arginine-rich (SR) protein family play an important role in both constitutive and regulated splicing of precursor mRNAs. Phosphorylation of the arginine/serine dipeptide-rich domain (RS domain) can modulate the activity and the subcellular localization of SR proteins. However, whether the SR protein family members are individually regulated and how this is achieved remain unclear. In this report we show that 5,6-dichloro-1 beta-D-ribofuranosyl-benzimidazole (DRB), an inhibitor of RNA polymerase II-dependent transcription, specifically induced hyperphosphorylation of SRp55 but not that of any other SR proteins tested. Hyperphosphorylation of SRp55 occurs at the RS domain and appears to require the RNA-binding activity. Upon DRB treatment, hyperphosphorylated SRp55 relocates to enlarged nuclear speckles. Intriguingly, SRp55 is specifically targeted for degradation by the proteasome upon overexpression of the SR protein kinase Clk/Sty. Although a destabilization signal is mapped within the C-terminal 43-amino acid segment of SRp55, its adjacent lysine/serine-rich RS domain is nevertheless critical for the Clk/Sty-mediated degradation. We report for the first time that SRp55 can be hyperphosphorylated under different circumstances whereby its fate is differentially influenced.

Animals↗

PubMed2003

A novel splicing regulator shares a nuclear import pathway with SR proteins.

Alternative splicing of precursor mRNA is often regulated by serine/arginine-rich proteins (SR proteins) and hnRNPs, and varying their concentration in the nucleus can be a mechanism for controlling splice site selection. To understand the nucleocytoplasmic transport mechanism of splicing regulators is of key importance. SR proteins are delivered to the nucleus by transportin-SRs (TRN-SRs), importin beta-like nuclear transporters. Here we identify and characterize a non-SR protein, RNA-binding motif protein 4 (RBM4), as a novel substrate of TRN-SR2. TRN-SR2 interacts specifically with RBM4 in a Ran-sensitive manner. TRN-SR2 indeed mediates the nuclear import of a recombinant protein containing the RBM4 C-terminal domain. This domain serves as a signal for both nuclear import and export, and for nuclear speckle targeting. Finally, both in vivo and in vitro splicing analyses demonstrate that RBM4 not only modulates alternative pre-mRNA splicing but also acts antagonistically to authentic SR proteins in splice site and exon selection. Thus, a novel splicing regulator with opposite activities to SR proteins shares an identical import pathway with SR proteins to the nucleus.

Active Transport, Cell Nucleus↗

PubMed2003

Nuclear Pnn/DRS protein binds to spliced mRNPs and participates in mRNA processing and export via interaction with RNPS1.

Pnn/DRS protein is associated with desmosomes and colocalizes with splicing factors in nuclear speckled domains. The potential interaction of Pnn with RNPS1, a pre-mRNA splicing factor and a component of the exon-exon junction complex, prompted us to examine whether Pnn is involved in nuclear mRNA processing. By immunoprecipitation, we found that Pnn associates preferentially with mRNAs produced by splicing in vitro. Oligonucleotide-directed RNase H digestion revealed that Pnn binds to the spliced mRNAs at a position immediately upstream of the splice junction and that 5' splice site utilization determines the location of Pnn in alternatively spliced mRNAs. Immunoprecipitation further showed that Pnn binds to mRNAs produced from a transiently expressed reporter in vivo. Although associated with mRNPs, Pnn is a nuclear-restricted protein as revealed by the heterokaryon assay. Overexpression of an amino-terminal fragment of Pnn that directly interacts with RNPS1 leads to blockage of pre-mRNA splicing. However, although suppression of Pnn expression shows no significant effect on splicing, it leads to some extent to nuclear accumulation of bulk poly(A)(+) RNA. Therefore, Pnn may participate, via its interaction with RNPS1, in mRNA metabolism in the nucleus, including mRNA splicing and export.

Biological Transport↗