The type-1 ribosome-inactivating protein OsRIP1 triggers caspase-independent apoptotic-like death in HeLa cells

Ribosome-inactivating proteins (RIPs) are capable of removing a specific adenine from 285 ribosomal RNA, thus inhibiting protein biosynthesis in an irreversible manner. In this study, recombinant OsRIP1, a type 1 RIP from rice (Oryza saliva L.), was investigated for its anti-proliferative properties. Human cervical cancer HeLa cells were incubated in the presence of OsRIP1 for 24-72 h. OsRIP1 treatment yielded an anti-proliferation response of the HeLa cells and resulted in apoptotic-like blebbing of the plasma membrane without causing DNA fragmentation. OsRIP1 labeled with FITC accumulated at the cell surface. Pull-down assays identified ASPP1 (Apoptosis-Stimulating Protein of p53 1) and IFITM3 (interferon-induced transmembrane protein 3) as potential interaction partners for OsRIP1. Transcript levels for several critical genes related to different signaling pathways were quantified by RT-qPCR. OsRIP1 provoked HeLa cells to undergo caspase-independent cell death, associated with a significant transcriptional upregulation of the apoptotic gene PUMA, interferon regulatory factor 1 (IRF1) and the autophagy-related marker LC3. No changes in caspase activities were observed. Together, these data suggest that apoptotic-like events were involved in OsRIP1-driven caspase-independent cell death that might trigger the IRF1 signaling pathway and LC3-mediated autophagy

A Diocleinae type II lectin from Dioclea lasiophylla Mart. Ex Benth seeds specific to alpha-lactose/GalNAc

A type II lectin, designated as DlyL2, was purified from Dioclea lasiophylla Mart ex Benth seeds and some of its physicochemical properties determined. The lectin demonstrated specificity for alpha-lactose and N-acetyl-D-galactosamine and was able to interact with porcine stomach mucin. DlyL2 has 0.78 % carbohydrates in its composition, therefore can be considered a glycoprotein. In addition, its hemagglutinating activity remained stable M a temperature of 90 degrees C and in a pH range from 5 to 10. Metal chelation treatment did not affect DlyL2 activity suggesting ifs not a metalloprotein. Dly12 showed an apparent mass of 31 kDa and average molecular mass of 26.371 kDa. Primary structure data could be generated from the partial amino acid sequence of DlyL2, with about 192 residues sequenced by a combination of Edman degradation and tandem mass spectrometry. The lectin is similar to other type II lectins from Diocleinae subtribe, as well as lectins derived from species of more ancient tribes of the Fabaceae family. In addition, DlyL2 exhibited no toxicity to Anemia sp. nauplii. The present study expands the knowledge about the specificity, structural and physicochemical properties of Diocleinae type II lectins