Functional of nucleolar AAA-ATPase NVL2 in ribosome biogenesis.

NVL2是调节蛋白复合物功能的aaa - atp酶家族的一员。研究表明NVL2与核糖体蛋白L5和RNA解旋酶DOB1相互作用，参与60S核糖体的生物发生。为了研究NVL2在核糖体生物发生中的作用，我们进行了以下研究。1. 已经证明，表达NVL2显性阴性突变体会损害60S核糖体的生物发生。为了研究这些细胞中rRNA前体的加工是否受损，我们进行了Northern印迹和脉冲追踪分析。这些实验没有显示出对rRNA前体加工的明显影响。这些结果表明，在完成rRNA前体的加工后，NVL2参与了核糖体生物发生的后期步骤。2. 利用培养细胞核提取物的蔗糖梯度分离，分析核糖体与NVL2或DOB1的相互作用。结果表明，NVL2与细胞核内的预核糖体相互作用。相反，在与核糖体相同的组分中没有观察到DOB，并且存在于低分子量组分中。然而，在显性阴性NVL2表达的细胞中，部分DOB1出现在60s前核糖体的部分中。这些结果表明NVL2可能作为分子伴侣，调节DOB1和前体之间的相互作用。

NVL2 is a member of the AAA-ATPase family, which regulates functions of protein complexes. It has been shown that NVL2 interacts with ribosomal protein L5 and RNA helicase DOB1 and is involved in the biogenesis of 60S ribosome. To investigate the role of NVL2 in ribosome biogenesis, following studies have been done. 1. It has been demonstrated that biogenesis of 60S ribosome is impaired by expressing a dominant negative mutant of NVL2. To investigate whether processing of rRNA precursor is impaired in these cells, Northern blotting and pulse-chase analysis were performed. These experiments did not show distinct effect on the processing of rRNA precursor. These results suggest that NVL2 is involved in a late step of ribosome biogenesis after the completion of the processing of rRNA precursors. 2. Interactions between preribosome and NVL2 or DOB1 were analyzed by using sucrose gradient fractionation of nuclear extracts from cultured cells. The results demonstrated that NVL2 interacts with preribosome in the nucleus. On the contrary, DOB was not observed in the same fractions with preribosome and existed in the low molecular weight fractions. However, in cells in which the dominant negative NVL2 is expressed, a part of DOB1 was appeared in the fractions of pre-60S ribosomes. These results suggest that NVL2 might act as a molecular chaperone, which regulates interaction between DOB1 and preribosome.