Integration and expression of 3′-truncated derivatives of the Neurospora crassa cyt-21 + gene, encoding a mitochondrial ribosomal protein, in Neurospora transformants

Abstract

We showed previously that the Neurospora crassa mutation cyt-21-1 causes abnormal splicing of cyt-21 pre-mRNA, putatively leading to an internally deleted mitochondrial ribosomal protein. In this study, deletion derivatives of the Neurospora crassa cyt-21+ gene are used to complement cyt-21-1. Large parts of the 3′ end of the gene, comprising almost the entire last exon, can be deleted without affecting the transformation frequency. Analysis of DNA from transformants shows that the transforming DNA integrates non-specifically into the genome of the host. Northern analysis showed that the 3′ end of the cyt-21+ gene can be altered dramatically without affecting the 3′ processing of transcripts. One deletion derivative, in which part of the coding region of the gene is removed, yields transformants expressing a carboxyterminally modified ribosomal protein. The direct relationship between in vitro modification of the cloned Neurospora cyt-21+ gene and the detection of a modified mitoribosomal protein in cyt-21 transformants provides firm evidence that the cyt-21+ gene encodes a mitoribosomal protein.