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Biomedical subjects

Weidong Hu

Publications and source records attributed to Weidong Hu.

11 recordsLinked to original sources
PubMed2026

Targeting the METTL1/m7G axis as a therapeutic strategy in myeloid leukemia.

N7-methylguanosine (m7G), a prevalent modification in transfer RNAs (tRNAs), is primarily catalyzed by the methyltransferase METTL1. Although growing evidence supports a role for METTL1 in various tumors, its therapeutic potential and precise function in leukemia stem cell (LSC) homeostasis remain largely unexplored. Here, we identify METTL1 as a key regulator of LSC self-renewal and homing within bone marrow (BM) microenvironment through catalyzing m7G formation on a specific tRNA, tRNAPheGAA, thereby promoting leukemogenesis. Mechanistically, METTL1 loss significantly reduces m7G abundance and steady-state levels of tRNAPheGAA, leading to translation suppression and degradation of transcripts enriched with tRNAPheGAA-related codons, such as hematopoietic cell kinase (HCK). Decreased HCK expression disrupts CXCR4 signaling, impairing LSC self-renewal and BM homing. Therapeutically, we characterized a small-molecule METTL1 inhibitor (M1i; NSC137443), through high-throughput screening. Pharmacological inhibition of METTL1 demonstrated potent antitumor efficacy by reducing tRNA m7G levels and disrupting the tRNAPheGAA/HCK/CXCR4 cascade. Notably, targeting METTL1 significantly reduces LSC frequency, delays leukemogenesis, and prolongs survival in multiple acute myeloid leukemia models. Together, our findings establish a previously unrecognized role for METTL1 and its target tRNAPheGAA in LSC homeostasis and provide compelling proof-of-concept evidence that METTL1 is a druggable epitranscriptomic target for antileukemia therapy.

Humans↗
PubMed2006

Mutation analysis of the short cytoplasmic domain of the cell-cell adhesion molecule CEACAM1 identifies residues that orchestrate actin binding and lumen formation.

CEACAM1-4S (carcinoembryonic antigen cell adhesion molecule 1, with 4 ectodomains and a short, 12-14 amino acid cytoplasmic domain) mediates lumen formation via an apoptotic and cytoskeletal reorganization mechanism when mammary epithelial cells are grown in a three-dimensional model of mammary morphogenesis. We show by quantitative yeast two-hybrid, BIAcore, NMR HSQC and STD, and confocal analyses that amino acids phenylalanine (Phe(454)) and lysine (Lys(456)) are key residues that interact with actin orchestrating the cytoskeletal reorganization. A CEACAM1 membrane model based on vitamin D-binding protein that predicts an interaction of Phe(454) at subdomain 3 of actin was supported by inhibition of binding of actin to vitamin D-binding protein by the cytoplasmic domain peptide. We also show that residues Thr(457) and/or Ser(459) are phosphorylated in CEACAM1-transfected cells grown in three-dimensional culture and that mutation analysis of these residues (T457A/S459A) or F454A blocks lumen formation. These studies demonstrate that a short cytoplasmic domain membrane receptor can directly mediate substantial intracellular signaling.

Actins↗
PubMed2006

The lipid-associated 3D structure of SPA, a broad-spectrum neuropeptide antagonist with anticancer properties.

[D-Arg(1), D-Trp(5,7,9), Leu(11)] substance P (SPA) belongs to a family of peptides including antagonist G and SpD that act as broad-spectrum neuropeptide antagonists at several peripheral receptors. The lipid-induced structure of these peptides may be important for the receptor interactions of these analogs. Thus we describe the tertiary structure of SPA in the presence of sodium dodecylsulfate micelles at pH 5.0, and 25 degrees C as determined from two-dimensional (1)H-NMR data recorded at 500 MHz. The resulting three-dimensional structure can be generally described as two type IV nonstandard turns around Arg(1)*, Pro(2), Lys(3), and Pro(4) and Gln(6), Trp(7)*, Phe(8), and Trp(9)* residues, respectively, inserted into the interfacial region of the micelles (the asterisks denote D-form amino acid). These turns juxtapose the N- and C-termini of SPA and may form the basis of this peptide's unique ability to inhibit peptide receptor interactions at multiple receptor types.

Antineoplastic Agents↗
PubMed2005

Small ubiquitin-like modifier (SUMO) recognition of a SUMO binding motif: a reversal of the bound orientation.

Sumoylation has recently been identified as an important mechanism that regulates protein interactions and localization in essential cellular functions, such as gene transcription, subnuclear structure formation, viral infection, and cell cycle progression. A SUMO binding amino acid sequence motif (SBM), which recognizes the SUMO moiety of modified proteins in sumoylation-dependent cellular functions, has been consistently identified by several recent studies. To understand the mechanism of SUMO recognition by the SBM, we have solved the solution structure of SUMO-1 in complex with a peptide containing the SBM derived from the protein PIASX (KVDVIDLTIESSSDEEEDPPAKR). Surprisingly, the structure reveals that the bound orientation of the SBM can reverse depending on the sequence context. The structure also reveals a novel mechanism of recognizing target sequences by a ubiquitin-like module. Unlike ubiquitin binding motifs, which all form helices and bind to the main beta-sheet of ubiquitin, the SBM forms an extended structure that binds between the alpha-helix and a beta-strand of SUMO-1. This study provides a clear mechanism of the SBM sequence variations and its recognition of the SUMO moiety in sumoylated proteins.

Amino Acid Motifs↗
PubMed2005

A solid-phase approach to novel purine and nucleoside analogs.

This paper describes a method for the preparation of purine analogs using the solid-phase approach. Nucleoside bases were constructed on Merrifield resin by sequential displacement of purine dichloride with amines, and after detachment, the purine analogs were condensed with d,l-ribofuranoside compounds by the Vorbrüggen method. Thereof, l-ribofuranoside was prepared from l-arabinose via the selective oxidation-reduction procedure of the 2-OH group. Some compounds exhibited moderate activity against HIV-1 in PBM cells.

Anti-HIV Agents↗
PubMed2004

Retention of conformational flexibility in HIV-1 Rev-RNA complexes.

Sequence-specific recognition between HIV-1 Rev and viral RNA mediates the nuclear export of the viral mRNA and thus is important for the viral life cycle. HIV Rev binds to its viral RNA target with high affinity and specificity and also binds to an in vitro selected RNA aptamer that has a significantly different sequence from the viral RNA target with a 6-fold higher affinity than its natural target. The high-resolution structures of HIV Rev Arg-rich motif (ARM) in complexes with the wild-type RNA and the RNA aptamer reveal that, despite the significantly different RNA sequences, the two complexes share similar structural features and the protein-RNA interactions are mediated mostly by the Arg side chains in Rev ARM. To gain further insight into the role of these Arg side chains in the sequence-specific protein-RNA recognition, we have characterized the flexibility of these Arg side chains at the interfaces of the two high-affinity complexes using (15)N R(1), R(2), nuclear Overhauser effect, and chemical-shift anisotropy dipolar cross-correlation relaxation measurements. The ARM peptide contains uniformly (13)C/(15)N-labeled Arg residues, and the RNA samples were unlabeled. Despite the apparently similar roles of Arg side chains in both complexes, most of them display a different dynamic behavior in the context of different RNA molecules, and extensive and highly diverse motions have been observed for all of these side chains that interact with RNA. Most of the differences in side-chain dynamics between the complexes cannot be inferred from the three-dimensional structures. Additionally, more than half of the residues have increased flexibility in the Rev-RNA aptamer complex that has a higher affinity. This study provides new insights into ARM-RNA recognition and indicates that retention of conformational flexibility is likely important in high-affinity ARM-RNA recognition.

Arginine↗
PubMed2004

Solution structure of the C-terminal domain of Ku80 suggests important sites for protein-protein interactions.

The solution structure of Ku80 CTD from residue 566 to 732 has been solved in order to gain insights into the mechanisms of its interactions with other proteins. The structure reveals a topology similar to several common scaffolds for protein-protein interactions, in the absence of significant sequence similarity to these proteins. Conserved surface amino acid residues are clustered on two main surface areas, which are likely involved in mediating interactions between Ku80 and other proteins. The Ku70/Ku80 heterodimer has been shown to be involved in at least three processes, nonhomologous end joining, transcription, and telomere maintenance, and thus it needs to interact with different proteins involved in these different processes. The three-dimensional structure of the Ku80 C-terminal domain and the availability of NMR chemical shift assignments provide a basis for further investigation of the interactions between Ku80 and other proteins in these Ku-dependent cellular functions.

Alkyl and Aryl Transferases↗
PubMed2003

A high sensitivity 3D experiment for measuring Calpha-Halpha residual dipolar coupling constants.

A new sensitivity improved approach is presented to measure the Calpha-Halpha scalar and dipolar coupling constants in 13C/15N-labeled proteins using a HA(CA)CONH scheme. The proposed experiment has significantly higher sensitivity than the previously published (HA)CA(CO)NH sequence, and provides accurate and straightforward measurements of the scalar and residual dipolar coupling constants. The sequence is easy to implement, and has been demonstrated on the C-terminal domain of the human Ku-80 protein (152 amino acid residues). On average, sensitivity is improved by 40% for both isotropic and anisotropic samples. The sensitivity enhancement is more pronounced for structured regions than unstructured regions, with an average of 50-60% enhancement being observed in the well-structured regions of the protein.

Algorithms↗
PubMed2002

Concerted motions in HIV-1 TAR RNA may allow access to bound state conformations: RNA dynamics from NMR residual dipolar couplings.

Ground-state dynamics in RNA is a critical precursor for structural adaptation observed ubiquitously in protein-RNA recognition. A tertiary conformational analysis of the stem-loop structural element in the transactivation response element (TAR) from human immunodeficiency virus type 1 (HIV-I) RNA is presented using recently introduced NMR methods that rely on the measurement of residual dipolar couplings (RDC) in partially oriented systems. Order matrix analysis of RDC data provides evidence for inter-helical motions that are of amplitude 46(+/-4) degrees, of random directional character, and that are executed about an average conformation with an inter-helical angle between 44 degrees and 54 degrees. The generated ensemble of TAR conformations have different organizations of functional groups responsible for interaction with the trans-activator protein Tat, including conformations similar to the previously characterized bound-state conformation. These results demonstrate the utility of RDC-NMR for simultaneously characterizing RNA tertiary dynamics and average conformation, and indicate an avenue for TAR complex formation involving tertiary structure capture.

Gene Products, tat↗
PubMed2002

Ribosome-associated factor Y adopts a fold resembling a double-stranded RNA binding domain scaffold.

Escherichia coli protein Y (pY) binds to the small ribosomal subunit and stabilizes ribosomes against dissociation when bacteria experience environmental stress. pY inhibits translation in vitro, most probably by interfering with the binding of the aminoacyl-tRNA to the ribosomal A site. Such a translational arrest may mediate overall adaptation of cells to environmental conditions. We have determined the 3D solution structure of a 112-residue pY and have studied its backbone dynamic by NMR spectroscopy. The structure has a betaalphabetabetabetaalpha topology and represents a compact two-layered sandwich of two nearly parallel alpha helices packed against the same side of a four-stranded beta sheet. The 23 C-terminal residues of the protein are disordered. Long-range angular constraints provided by residual dipolar coupling data proved critical for precisely defining the position of helix 1. Our data establish that the C-terminal region of helix 1 and the loop linking this helix with strand beta2 show significant conformational exchange in the ms- micro s time scale, which may have relevance to the interaction of pY with ribosomal subunits. Distribution of the conserved residues on the protein surface highlights a positively charged region towards the C-terminal segments of both alpha helices, which most probably constitutes an RNA binding site. The observed betaalphabetabetabetaalpha topology of pY resembles the alphabetabetabetaalpha topology of double-stranded RNA-binding domains, despite limited sequence similarity. It appears probable that functional properties of pY are not identical to those of dsRBDs, as the postulated RNA-binding site in pY does not coincide with the RNA-binding surface of the dsRBDs.

Amino Acid Sequence↗