Abstract
Regulatory circuits have been defined in Neurospora cmsa within which multiple unlinked genes are controlled in a positive fashion by major regulatory genes. It has been suggested that repetitive DNA sequences situated adjacent to structural genes may serve as regulatory sites. Experiments are presented which examine the amount and organization of the repetitive sequences of Neurospora. The genome consists of 2.7 X 10’ base pairs and is composed of 2% foldback, 8% repetitive, and 90% single copy sequences. Our results indicate that the repeated DNA sequences are organized in stretches of 10,000 base pairs or longer. Experiments using pure ribosomal DNA isolated from cloned recombinant DNA demonstrated that rDNA represents 7% of total nuclear DNA and 88% of the repetitive component. The rDNA sequence is repeated about 185 times per nucleus. Since the ribosomal DNA units are believed to be tandemly repeated many times, their clustering can largely account for the long repetitive stretches and their lack of interspersion with single copy DNA. These results argue that repetitive DNA sequences identified by renaturation experiments do not serve a regulatory function in Neurospora. About 350 foci of inverted repeat, or foldback, DNA sequences occur in the genome and are interspersed with single copy DNA. The relatively small number of foldback foci suggest that inverted repeats also cannot be the primary recognition elements for control of gene expression in Neurospora. We suggest that very short repeated sequences, undetected by renaturation experiments, may serve as recognition sites for eukaryotic gene regulation.
Highlights
Regulatorycircuits have been defined in Neu- strated for phosphorus metabolism [2],sulfur metabolism [5], rospora c m s a within which multiple unlinked genes and synthesis of branched chain amino acids [6]
DNA sequences identified byrenaturation experiments quence organization in most of the organisms examined (10, d o not serve a regulatory functionin Neurospora.About 11).This patternis typified by short repetitive sequences, 200
SequeNniecnuersospora organized in a short period interspersion pattern [19].In this of Rigby et al [25].A 200-1.11reaction mixture containing 50 mM phosphate buffer (PB), paper, we present experimenths at characterizethe nature of pH 7.6,5 m~ MgClz,15p~ 5”dGTP, 5’-dCTP, and 5”dATP, 250 pCi the repetitive DNA sequences of Neurospora and their organizationin the genome
Summary
It has been suggested that repetitive DNA se- olism, are clustered and regulated by a closely linked control quences situated adjacent to structural genes m a y gene [7,8]. these three qa genes are tightly linked, serve as regulatory sites Experiments are presented they aretranscribed as separateunits. Experiments using pure ribosomal DNA isosesses a similar adjacent DNA sequence which serves as a recognition site for a common regulatory protein [9] Such considerations suggest that each controlcircuit should contain a family of repeated DNA sequences, the degree of repetition corresponding to the number of structural genes within the lated from cloned recombinant DNA demonstratetdhat circuit. These duplexes are proposed to regulate the turnover and processing of nuclear transcripts into functional mRNA’s
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