Russell L. Malmberg

A standardized kinesin nomenclature

In recent years the kinesin superfamily has become so large that several different naming schemes have emerged, leading to confusion and miscommunication. Here, we set forth a standardized kinesin nomenclature based on 14 family designations. The scheme unifies all previous phylogenies and nomenclature proposals, while allowing individual sequence names to remain the same, and for expansion to occur as new sequences are discovered.

Analysis of septins across kingdoms reveals orthology and new motifs.

BackgroundSeptins are cytoskeletal GTPase proteins first discovered in the fungus Saccharomyces cerevisiae where they organize the septum and link nuclear division with cell division. More recently septins have been found in animals where they are important in processes ranging from actin and microtubule organization to embryonic patterning and where defects in septins have been implicated in human disease. Previous studies suggested that many animal septins fell into independent evolutionary groups, confounding cross-kingdom comparison.ResultsIn the current work, we identified 162 septins from fungi, microsporidia and animals and analyzed their phylogenetic relationships. There was support for five groups of septins with orthology between kingdoms. Group 1 (which includes S. cerevisiae Cdc10p and human Sept9) and Group 2 (which includes S. cerevisiae Cdc3p and human Sept7) contain sequences from fungi and animals. Group 3 (which includes S. cerevisiae Cdc11p) and Group 4 (which includes S. cerevisiae Cdc12p) contain sequences from fungi and microsporidia. Group 5 (which includes Aspergillus nidulans AspE) contains sequences from filamentous fungi. We suggest a modified nomenclature based on these phylogenetic relationships. Comparative sequence alignments revealed septin derivatives of already known G1, G3 and G4 GTPase motifs, four new motifs from two to twelve amino acids long and six conserved single amino acid positions. One of these new motifs is septin-specific and several are group specific.ConclusionOur studies provide an evolutionary history for this important family of proteins and a framework and consistent nomenclature for comparison of septin orthologs across kingdoms.

Regulation of Arabidopsis thaliana (L.) Heynh Arginine Decarboxylase by Potassium Deficiency Stress

Arginine decarboxylase (ARGdc) is the first enzyme in one of the two pathways to putrescine in plants. ARGdc enzyme activity has been shown to be induced by many environmental factors, including potassium deficiency stress. We investigated the mechanism for induction of ARGdc activity during potassium deficiency stress in Arabidopsis thaliana (L.) Heynh. We show that A. thaliana responds to potassium deficiency stress by increasing ARGdc activity by up to 10-fold over unstressed plants with a corresponding increase in putrescine levels of up to 20-fold. Spermidine and spermine levels do not increase proportionately. Northern analysis showed no increase in ARGdc mRNA levels correlated with the increase in ARGdc enzyme activity. Western analysis revealed that there was no difference between ARGdc protein levels in stressed plants compared with controls. The increase in ARGdc enzyme activity due to potassium deficiency stress does not appear to involve changes in mRNA or protein abundance.

Epistasis for fitness-related quantitative traits in Arabidopsis thaliana grown in the field and in the greenhouse

The extent to which epistasis contributes to adaptation, population differentiation, and speciation is a long-standing and important problem in evolutionary genetics. Using recombinant inbred (RI) lines of Arabidopsis thaliana grown under natural field conditions, we have examined the genetic architecture of fitness-correlated traits with respect to epistasis; we identified both single-locus additive and two-locus epistatic QTL for natural variation in fruit number, germination, and seed length and width. For fruit number, we found seven significant epistatic interactions, but only two additive QTL. For seed germination, length, and width, there were from two to four additive QTL and from five to eight epistatic interactions. The epistatic interactions were both positive and negative. In each case, the magnitude of the epistatic effects was roughly double that of the effects of the additive QTL, varying from −41% to +29% for fruit number and from −5% to +4% for seed germination, length, and width. A number of the QTL that we describe participate in more than one epistatic interaction, and some loci identified as additive also may participate in an epistatic interaction; the genetic architecture for fitness traits may be a network of additive and epistatic effects. We compared the map positions of the additive and epistatic QTL for germination, seed width, and seed length from plants grown in both the field and the greenhouse. While the total number of significant additive and epistatic QTL was similar under the two growth conditions, the map locations were largely different. We found a small number of significant epistatic QTL × environment effects when we tested directly for them. Our results support the idea that epistatic interactions are an important part of natural genetic variation and reinforce the need for caution in comparing results from greenhouse-grown and field-grown plants.

Analysis of a cDNA encoding arginine decarboxylase from oat reveals similarity to the Escherichia coli arginine decarboxylase and evidence of protein processing

SummaryArginine decarboxylase is the first enzyme in one of the two pathways of putrescine synthesis in plants. We purified arginine decarboxylase from oat leaves, obtained N-terminal amino acid sequence, and then used this information to isolate a cDNA encoding oat arginine decarboxylase. Comparison of the derived amino acid sequence with that of the arginine decarboxylase gene from Escherichia coli reveals several regions of sequence similarity which may play a role in enzyme function. The open reading frame (ORF) in the oat cDNA encodes a 66 kDa protein, but the arginine decarboxylase polypeptide that we purified has an apparent molecular weight of 24 kDa and is encoded in the carboxyl-terminal region of the ORF. A portion of the cDNA encoding this region was expressed in E. coli, and a polyclonal antibody was developed against the expressed polypeptide. The antibody detects 34 kDa and 24 kDa polypeptides on Western blots of oat leaf samples. Maturation of arginine decarboxylase in oats appears to include processing of a precursor protein.

Maximum Likelihood Methods Reveal Conservation of Function Among Closely Related Kinesin Families

Abstract. We have reconstructed the evolution of the anciently derived kinesin superfamily using various alignment and tree-building methods. In addition to classifying previously described kinesins from protists, fungi, and animals, we analyzed a variety of kinesin sequences from the plant kingdom including 12 from Zea mays and 29 from Arabidopsis thaliana. Also included in our data set were four sequences from the anciently diverged amitochondriate protist Giardia lamblia. The overall topology of the best tree we found is more likely than previously reported topologies and allows us to make the following new observations: (1) kinesins involved in chromosome movement including MCAK, chromokinesin, and CENP-E may be descended from a single ancestor; (2) kinesins that form complex oligomers are limited to a monophyletic group of families; (3) kinesins that crosslink antiparallel microtubules at the spindle midzone including BIMC, MKLP, and CENP-E are closely related; (4) Drosophila NOD and human KID group with other characterized chromokinesins; and (5) Saccharomyces SMY1 groups with kinesin-I sequences, forming a family of kinesins capable of class V myosin interactions. In addition, we found that one monophyletic clade composed exclusively of sequences with a C-terminal motor domain contains all known minus end-directed kinesins.

QTL-based evidence for the role of epistasis in evolution

The extent to which epistasis contributes to adaptation and speciation has been a controversial topic in evolutionary genetics. One experimental approach to study epistasis is based on quantitative trait locus (QTL) mapping using molecular markers. Comparisons can be made among all possible pair-wise combinations of the markers, irrespective of whether an additive QTL is associated with a marker; several software packages have been developed that facilitate this. We review several examples of using this approach to identify epistatic QTLs for traits of evolutionary or ecological interest. While there is variability in the results, the number of epistatic QTL interactions is often greater than or equal to the number of additive QTLs. The magnitude of epistatic effects can be larger than the additive effects. Thus, epistatic interactions seem to be an important part of natural genetic variation. Future studies of epistatic QTLs could lead to descriptions of the genetic networks underlying variation for fitness-related traits.

Longitudinal trends in climate drive flowering time clines in North American Arabidopsis thaliana

Introduced species frequently show geographic differentiation, and when differentiation mirrors the ancestral range, it is often taken as evidence of adaptive evolution. The mouse-ear cress (Arabidopsis thaliana) was introduced to North America from Eurasia 150–200 years ago, providing an opportunity to study parallel adaptation in a genetic model organism. Here, we test for clinal variation in flowering time using 199 North American (NA) accessions of A. thaliana, and evaluate the contributions of major flowering time genes FRI, FLC, and PHYC as well as potential ecological mechanisms underlying differentiation. We find evidence for substantial within population genetic variation in quantitative traits and flowering time, and putatively adaptive longitudinal differentiation, despite low levels of variation at FRI, FLC, and PHYC and genome-wide reductions in population structure relative to Eurasian (EA) samples. The observed longitudinal cline in flowering time in North America is parallel to an EA cline, robust to the effects of population structure, and associated with geographic variation in winter precipitation and temperature. We detected major effects of FRI on quantitative traits associated with reproductive fitness, although the haplotype associated with higher fitness remains rare in North America. Collectively, our results suggest the evolution of parallel flowering time clines through novel genetic mechanisms.

Efficient Parameterized Algorithms for Biopolymer Structure-Sequence Alignment

Computational alignment of a biopolymer sequence (e.g., an RNA or a protein) to a structure is an effective approach to predict and search for the structure of new sequences. To identify the structure of remote homologs, the structure-sequence alignment has to consider not only sequence similarity, but also spatially conserved conformations caused by residue interactions and, consequently, is computationally intractable. It is difficult to cope with the inefficiency without compromising alignment accuracy, especially for structure search in genomes or large databases. This paper introduces a novel method and a parameterized algorithm for structure-sequence alignment. Both the structure and the sequence are represented as graphs, where, in general, the graph for a biopolymer structure has a naturally small tree width. The algorithm constructs an optimal alignment by finding in the sequence graph the maximum valued subgraph isomorphic to the structure graph. It has the computational time complexity O(k3N2) for the structure of N residues and its tree decomposition of width t. Parameter k, small in nature, is determined by a statistical cutoff for the correspondence between the structure and the sequence. This paper demonstrates a successful application of the algorithm to RNA structure search used for noncoding RNA identification. An application to protein threading is also discussed

Resolving phylogenetic relationships of the recently radiated carnivorous plant genus Sarracenia using target enrichment

The North American carnivorous pitcher plant genus Sarracenia (Sarraceniaceae) is a relatively young clade (<3 million years ago) displaying a wide range of morphological diversity in complex trapping structures. This recently radiated group is a promising system to examine the structural evolution and diversification of carnivorous plants; however, little is known regarding evolutionary relationships within the genus. Previous attempts at resolving the phylogeny have been unsuccessful, most likely due to few parsimony-informative sites compounded by incomplete lineage sorting. Here, we applied a target enrichment approach using multiple accessions to assess the relationships of Sarracenia species. This resulted in 199 nuclear genes from 75 accessions covering the putative 8-11 species and 8 subspecies/varieties. In addition, we recovered 42kb of plastome sequence from each accession to estimate a cpDNA-derived phylogeny. Unsurprisingly, the cpDNA had few parsimony-informative sites (0.5%) and provided little information on species relationships. In contrast, use of the targeted nuclear loci in concatenation and coalescent frameworks elucidated many relationships within Sarracenia even with high heterogeneity among gene trees. Results were largely consistent for both concatenation and coalescent approaches. The only major disagreement was with the placement of the purpurea complex. Moreover, results suggest an Appalachian massif biogeographic origin of the genus. Overall, this study highlights the utility of target enrichment using multiple accessions to resolve relationships in recently radiated taxa.