Site-directed mutagenesis of PsaA residue W693 affects phylloquinone binding and function in the photosystem I reaction center of Chlamydomonas reinhardtii

To investigate the environment of the phylloquinone secondary electron acceptor A(1) within the photosystem I reaction center, we have carried out site-directed mutagenesis of two tryptophan residues (W693 and W702) in the PsaA subunit of Chlamydomonas reinhardtii. One of these conserved tryptophans (W693) is predicted to be close to the phylloquinone and has been implicated in the interaction of A(1) with an aromatic residue through pi--pi stacking. We find that replacement of W702 with either histidine or leucine has no effect on the electronic structure of A(1)(*-) or on forward electron transfer from A(1)(*-) to the iron--sulfur center F(x). In contrast, the same mutations of W693 alter the electronic structure of the photoaccumulated A(1)(*-) and slow forward electron transfer as measured by the decay of the electron spin-polarized signal arising from the P700(*+)/A(1)(*-) radical pair. These results provide support for the hypothesis that W693 has a role in poising the redox potential of A(1)/A(1)(*-) so it can reduce F(x), and they indirectly provide evidence for electron transfer along the PsaA-side branch of cofactors in PSI.     

Efficient foreign gene expression in Chlamydomonas reinhardtii mediated by an endogenous intron

Heterologous genes introduced into the nuclear genome of Chlamydomonas reinhardtii are often poorly expressed. To understand the molecular mechanisms underlying this effect, we examined the influence of various factors on the expression of a chimeric transgene that confers resistance to zeomycin. This marker comprises the bacterial ble gene flanked by 5′ and 3′ sequences from the Chlamydomonas RBCS2 gene. We found that the frequency with which transformants are recovered is significantly increased when ble is fused to shorter versions of the RBCS2 promoter and when Chlamydomonas introns are introduced into the coding region of ble. The latter effect is particularly evident in the case of the first intron of RBCS2, which dramatically stimulates the transformation frequency and the level of ble expression. We found that this improvement is mediated in part by an enhancer element within the intron sequence, and that this element acts in an orientation-independent manner and is effective when placed either upstream or downstream of the promoter. Our results demonstrate that stable high-level expression of a foreign gene in Chlamydomonas is possible, and highlight a potential role of introns as modulators of gene expression in this alga.     

A lobster claw abnormality in the common marmoset (Callithrix jacchus)

A congenital abnormality of the digits, similar to the 'lobster claw' deformity of humans, was observed in 8 common marmosets. The condition varied in severity and in effect on the animals; it appeared to be due to a single autosomal recessive trait.     

Mitochondrial DNA differentiation during the speciation process in Peromyscus

We address the problem of the possible significance of biological speciation to the magnitude and pattern of divergence of asexually transmitted characters in bisexual species. The empirical data for this report consist of restriction endonuclease site variability in maternally transmitted mitochondrial DNA (mtDNA) isolated from 82 samples of Peromyscus polionotus and P. leucopus collected from major portions of the respective species' ranges. Data are analyzed together with previously published information on P. maniculatus, a sibling species to polionotus. Maps of restriction sites indicate that all of the variation observed can be reasonably attributed to base substitutions leading to loss or gain of particular recognition sites. Magnitude of mtDNA sequence divergence within polionotus (maximum approximately equal to 2%) is roughly comparable to that observed within any of five previously identified mtDNA assemblages in maniculatus. Sequence divergence within leucopus (maximum approximately equal to 4%) is somewhat greater than that within polionotus. Consideration of probable evolutionary links among mtDNA restriction site maps allowed estimation of matriarchal phylogenies within polionotus and leucopus. Clustering algorithms and qualitative Wagner procedures were used to generate phenograms and parsimony networks, respectively, for the between-species comparisons. Three simple graphical models are presented to illustrate some conceivable relationships of mtDNA differentiation to speciation. In theoretical case I, each of two reproductively defined species (A and B) is monophyletic in matriarchal genealogy; the common female ancestor of either species can either predate or postdate the speciation. In case II, neither species is monophyletic in matriarchal genotype. In case III, species B is monophyletic but forms a subclade within A which is thus paraphyletic with respect to B. The empirical results for mtDNA in maniculatus and polionotus appear to conform closely to case III. These theoretical and empirical considerations raise a number of questions about the general relationship of the speciation process to the evolution of uniparentally transmitted traits. Some of these considerations are presented, and it is suggested that the distribution patterns of mtDNA sequence variation within and among extant species should be of considerable relevance to the particular demographies of speciation.