Daughter Cell Completion

F l g u r a 1.10 Morphological cha n g a a otoaarvad during

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the onset of differentiation however, all three processes are observed to show characteristic increases. RNA

synthesis begins within the first fifteen minutes of differentiation (Dow et al., 1983) and continues at a constant rate which is increased at about 3h (Potts & Dow,

1979).

The rate of protein synthesis appears to mirror that of RNA synthesis, the increase in rate at about 3h

corresponding with daughter cell synthesis (Dow et a l ., 1983). Qualitative differences between the soluble proteins of inhibited swarmer cells and reproductive cells have been observed by two dimensional gel electrophoresis. Changes in the synthesis of specific polypeptides have also been

observed during differentiation. Interestingly, a 12kD polypeptide which is subject to rapid turnover in the

inhibited swarmer cell is specifically degraded at the start of differentiation. This polypeptide may be associated with maintenance of swarmer cell inhibition (Dow et al., 1983).

DNA replication in differentiating swarmer cells is initiated at about 2h, coincidentally with prostheca

synthesis, and continues at a constant rate throughout the rest of the developmental sequence (Potts & Dow, 1979).

Although little is known about the control of gene expression during the differentiation of R. vannielii swarmer cells, work to date suggests several possibilities.

1. Nucleoid conformation

The nucleoid of the swarmer cell, which is compact and centrally located, undergoes a change in conformation after initiation of differentiation to became elongated and moves toward the pole at which the prostheca will form

40

(Whittenbury & Dow, 1977). This position is maintained until after daughter cell completion when the nucleoid returns to the centre of the cell but retains its extended conformation. These changes in nucleoid conformation may be involved in the transcription and replication of the swarmer cell genome.

2. RNA polymerase modification

Recent work has shown that although no qualitative changes can be found, quantitative changes in RNA polymerase core-associated polypeptides occur during differentiation. Specifically, a decrease in a 95kD polypeptide and an increase in a 70kD polypeptide are observed (Dow et al.,

1983). These changes may bring about a switch in

transcriptional specificity which stimulates expression of developmental genes.

3. Genome rearrangements

Rhodomicrobium vannielii possesses unusually large amounts of inverted repeat sequence DNA (Potts et.al.,

1980). Such sequences are ubiquitous in translocatable DNA elements (Calos & Miller, 1980j Kleckner, 1981) and may also have effects on DNA secondary structure (Lilley, 1980). It is possible that the unidirectional switch by which swarmer cells become reproductively active involves a rearrangement of specific DNA sequences. This will be considered further

4 1 1.3 Inverted Repeat Sequences

1.3.1 Introduction

In this section a -few of the structural and functional properties of inverted repeat sequences will be discussed.

Interest in this type of sequence stems from the ability of each of the strands of any inverted repeat sequence to anneal with itself to form a duplex region of DNA.

i . e. ABCDEF...fedcba abcdef... FEDCBA ABCDEF abcdef abcdef ABCDEF Inverted repeat sequences possess 180° rotational

symmetry, sometimes called twofold or dyad symmetry, about a point midway between the repeats. The non-repeated DNA between the repeats may be of any length and where the intervening gap is small (<10bp) the inverted repeat structure may be called a palindrome.

These sequences were originally observed as a very rapidly renaturing fraction of DNA from both prokaryotes and eukaryotes (Bernard!, 1962} Alberts & Doty, 1968; Walker & McLaren, 1969). This concentration- independent

renaturation was initially thought to be due to covalent cross-links between the strands of duplex DNA (Alberts fc Doty, 1968), but was later shown to be due to the presence of inverted repeat sequences (Wilson t> Thomas, 1974).

These sequences appear to be a ubiquitous feature of all DNAs and in some cases are present in quantities greater than predicted by statistical methods (Day t< Blake, 1982). This has led to the suggestion that inverted repeat

42

sequences have some -function or property which has allowed their conservation in evolution. Inverted repeat sequences are required in most DNA rearrangements and are involved in some mechanisms of control of gene expression. They are present at the ends of transposable and invertible elements; act as sites for protein recognition, initiation of DNA replication, termination of trancription and processing of RNA molecules; and may give rise to unusual DNA secondary structures. The presence of this class of sequences in bacteria which exhibit differentiation, in quantities

similar to that found in eukaryotes, suggests that the investigation of the function of inverted repeat sequences

in differentiating bacteria might yield valuable results. This will be discussed further in section 1.3.4.

1.3.2 Distribution and Abundance of Inverted Repeat Sequences

1.3.2.1 Eukaryotic inverted repeats

Inverted repeat sequences appear to be a general

feature of eukaryotic DNA. They have been found in the DNA of many organisms, their organelles, and in viruses. Wilson and Thomas (1974) determined that for Drosophila spp.. mouse and human DNA at least, inverted repeats average 300-1200bp in length and are arranged in small clusters interspersed throughout the genome. This distribution was confirmed by Schmid et al. (197S) who found inverted repeat sequences of up to ISkb interspersed every 4O-0Okb in Drosophila DNA. They estimated that a total of 2000-4000 Inverted repeat pairs are present in the Drosophi1 a genome, accounting for