Endonucleolytic cleavage of the coding region determinant (CRD) of c-myc mRNA appears to play a critical role in regulating c-myc mRNA turnover. Using ³²P-labeled c-myc CRD RNA as substrate, we have purified and identified two endoribonucleases from rat liver polysomes that are capable of cleaving the transcript in vitro. A 17-kDa enzyme was identified as RNase1. Apurinic/apyrimidinic (AP) DNA endonuclease 1 (APE1) was identified as the 35-kDa endoribonuclease that preferentially cleaves in between UA and CA dinucleotides of c-myc CRD RNA. APE1 was further confirmed to be the 35-kDa endoribonuclease because: (i) the endoribonuclease activity of the purified 35-kDa native enzyme was specifically immuno-depleted with APE1 monoclonal antibody, and (ii) recombinant human APE1 generated identical RNA cleavage patterns as the native liver enzyme. Studies using E96A and H309N mutants of APE1 suggest that the endoribonuclease activity for c-myc CRD RNA shares the same active center with the AP-DNA endonuclease activity. Transient knockdown of APE1 in HeLa cells led to increased steady-state level of c-myc mRNA and its half-life. We conclude that the ability to cleave RNA dinucleotides is a previously unidentified function of APE1 and it can regulate c-myc mRNA level possibly via its endoribonuclease activity.

The c-myc mRNA coding region determinant-binding protein (CRD-BP) has high affinity for the coding region determinant (CRD) of c-myc mRNA. Such affinity is believed to protect c-myc CRD from endonucleolytic attack. We have recently purified a mammalian endoribonuclease which can cleave within the c-myc CRD in vitro. The availability of this purified endonuclease has made it possible to directly test the interaction between CRD-BP and the endonuclease in regulating c-myc CRD RNA cleavage. In this study, we have identified the coding region of MDR-1 RNA as a new target for CRD-BP. CRD-BP has the same affinity for c-myc CRD nts 1705–1886 and MDR-1 RNA nts 746–962 with K_d of 500 nM. The concentration-dependent affinity of CRD-BP to these transcripts correlated with the concentration-dependent blocking of endonuclease-mediated cleavage by CRD-BP. In contrast, three other recombinant proteins tested which had no affinity for c-myc CRD did not block endonuclease-mediated cleavage. Finally, we have identified RNA sequences required for CRD-BP binding. These results provide the first direct evidence that CRD-BP can indeed protect c-myc CRD cleavage initiated by an endoribonuclease, and the framework for further investigation into the interactions between CRD-BP, c-myc mRNA, MDR-1 mRNA and the endoribonuclease in cells.

Apurinic/apyrimidinic endonuclease 1 (APE1), an essential protein in mammals, is known to be involved in base excision DNA repair, acting as the major abasic endonuclease; the protein also functions as a redox coactivator of several transcription factors that regulate gene expression. Recent findings highlight a novel role for APE1 in RNA metabolism. The new findings are as follows: (i) APE1 interacts with rRNA and ribosome processing protein NPM1 within the nucleolus; (ii) APE1 interacts with proteins involved in ribosome assembly (i.e., RLA0, RSSA) and RNA maturation (i.e., PRP19, MEP50) within the cytoplasm; (iii) APE1 cleaves abasic RNA; and (iv) APE1 cleaves a specific coding region of c-myc mRNA in vitro and influences c-myc mRNA level and half-life in cells. Such findings on the role of APE1 in the posttranscriptional control of gene expression could explain its ability to influence diverse biological processes and its relocalization to cytoplasmic compartments in some tissues and tumors. In addition, we propose that APE1 serves as a "cleansing" factor for oxidatively damaged abasic RNA, establishing a novel connection between DNA and RNA surveillance mechanisms. In this review, we introduce questions and speculations concerning the role of APE1 in RNA metabolism and discuss the implications of these findings in a broader evolutionary context.

Mushrooms, the fruiting bodies of fungi, are known for a long time in different cultures around the world to possess medicinal properties and are used to treat various human diseases. Mushrooms that are parts of traditional medicine in Asia had been extensively studied and this has led to identification of their bioactive ingredients. North America, while home to one of the world’s largest and diverse ecological systems, has not subjected its natural resources especially its diverse array of mushroom species for bioprospecting purposes: Are mushrooms native to North America a good source for drug discovery? In this review, we compile all the published studies up to September 2020 on the bioprospecting of North American mushrooms. Out of the 79 species that have been investigated for medicinal properties, 48 species (60%) have bioactivities that have not been previously reported. For a mere 16 selected species, 17 new bioactive compounds (10 small molecules, six polysaccharides and one protein) have already been isolated. The results from our literature search suggest that mushrooms native to North America are indeed a good source for drug discovery.