Grade Assignment: Grades will be assigned based upon the percentage of total points accumulated according to the following scale: 100 - 92% = A; 91 - 88% = AB; 87 - 82% = B; 81 - 78% = BC; 77 - 72% = C; 71 - 68% = CD; 67 - 60% = D; below 59% = F
To determine your approximate grade at any time during the semester, simply divide the total number of points you have accumulated by the total possible. This information will always be provided. In addition, I will periodically provide you with a grade report. You should check this report for accuracy and to give you an indication of your progress. Keep all of your graded work, including lab work, for your records.
Never hesitate to come and talk to me about your grade, or any aspect of the course, at any time during the semester!
S/U grading can be requested at any time during the regular semester. You must submit your request in writing on a separate sheet of paper (not on a test or other assignment). Note: I do not recommend that anyone take the course for S/U grading, especially considering this is a majors course, but this is ultimately your decision. After final grades have been submitted to the Registrar, I will not change a letter grade to S/U or vice versa. Note: "S" is awarded for a letter grade of "C" and above.
Appeals: You have the option of appealing the grading you've received on any exam question (or assignment). To do so, type on a separate sheet of paper your rationale for why you should receive credit for the question. Be sure to frame your argument carefully and concisely. Turn in your typed appeal and the exam/assignment to me before the next exam - no appeal will be accepted after the date of the subsequent exam (or two weeks after an assignment is returned). Please note that if I have made any errors in grading your exams (i.e., incorrectly counted up points, mis-marked a question) please see me immediately and I will correct the error without the need for an appeal.
Email: I typically check and respond to email first thing in the morning (between 8 & 9:00 am) and before I leave in the evening (usually about 5:30 pm). If you send an email after about 5:00 pm I will not see it nor respond to it until the following day because I do not check email after leaving my office. Please plan accordingly.
Personal File, References And Cubby: You will have a file folder in a file box in our lab room. You may use it to store papers/notes/etc. In addition, I will place in this file any assignments not returned to you personally, extra copies (if any) of handouts, and course readings. When in doubt or need, check here. Copies of references cited in class will be placed in the Reference File that is also in the lab. You will also be assigned a "herbarium cubby" for storing plant specimens.
Honor Code: I run this class on the Honor Code system; in other words, I trust you to do your own work at all times. If you violate my trust, the consequences will be severe (first offense = failing exam/assignment; second offense = failing course). If you have even the slightest doubt that an activity violates the Honor Code - don't do it. For more information, consult our institutional policy on Academic Honesty.
Bonus Work: You will have the opportunity to earn bonus points by attending lectures, analyzing journal articles, participating in formal nature walks, or even reporting on science-related television programs. If it is "scientific" and can be reasonably considered to pertain to our course you can earn bonus points for participating in the activity. Obtain a "Bio-Bonus Form" and turn it in to me following the activity. Bonus work is due no later than Study Day. As a general guide - lectures are worth five bonus points, journal summaries and book reviews are worth 3 bonus points, and summaries of TV programs are worth 1 points. For other activities, we will assign an appropriate number of points. If in question, ask. The Bonus Card can be obtained online. As a rule of thumb, more than 20 bonus points will have little impact on your final grade.
Attendance:Each class I will pass around an attendance sheet to sign/initial. To reward you for your attendance you will receive a "cool sticker". You will not be penalized for missing class, but remember that being absent does not excuse you from completing assignments on time (i.e., turning in any that are due and getting the assignment for the next class). In general, you will not be able to make up anything missed in lecture or lab.
Classroom Etiquette: The following is a list of suggestions to make our classroom experience as enjoyable and productive as possible for all of us:
- Please turn off your mobile phone before class
- Obviously, if you need to use the toilet during class, please do so. However, please attempt to use the toilet before class so that you do not disrupt class proceedings when leaving/returning to your seat and you do not miss any class activities
- Unless you absolutely need to leave class before the period ends, please wait for me to dismiss the class before packing up your books, etc.
- If you bring food to class, please eat quietly and clean up after yourself
- Tidy your area before leaving class - return any supplies to the front desk, etc., throw away garbage, papers, etc.
- If you arrive late, please sit in the first available seat to minimize disrupting other students
- Please do not talk, pass notes, send text messages, etc during class.
Visitors: Visitors to our classroom are welcome. Please introduce your visitors to me. And, they should plan to participate (as best they can) in class activities.
Late Assignments: I expect that assignments will be turned in on time. I reserve the right to accept/refuse late assignments and to deduct points accordingly for any late assignments that are graded.
Pride:I believe that the appearance of an assignment is a reflection of the quality of the work and the degree of respect it deserves. Thus, for your benefit I require that: (1) Written assignments must be typed. There will be many obvious exceptions. For example, any assignments completed in your field notebook or assignment book need not be typed. If in question about whether an assignment should be typed, please ask. Assignments not typed will be penalized 50% of the total possible points; (2) Assignments with multiple pages must be stapled. Any assignment that is not stapled will automatically loose 2 points. Please note that I have a stapler available with me in every class; (3) Frayed edges - any assignment turned in on paper with frayed edges ripped out of a spiral bound notebook will automatically loose 2 points. If you use spiral notebooks that's fine - simply remove the edges before turning in the assignment.
Course/Teaching Philosophy: This is a mastery-based course. In other words, anyone can earn an "A" if he/she "masters" course materials. Since you will know exactly what material to master, there is nothing standing between you and an "A" except yourself.
I like to consider myself as a "coach" and you are one of the players. Among other things, the coach of a sports team must: (1) teach a player techniques to play better; (2) provide motivation for a player to achieve success; and (3) evaluate the skills of a player. Like coaches, a classroom teacher serves these same roles; to teach, provide motivation for learning and evaluating (grading) the success of student. Thus, we are on the same team trying to win the "plant taxonomy game". To help foster team spirit, we may occasionally do motivational cheers, listen to music and probably most importantly, we will work together cooperatively.
I also think that learning should be enjoyable. Hopefully we will laugh together and have fun. Stamps, stars and stickers will adorn some of your graded assignments. "Botanical" music will greet you when you arrive in class. This is all done in good fun, to make our learning environment more pleasant. Yet, we will always be respectful of one another. Some students in the past have commented that they think some of what we'll do is "childish." I hope so because I want to generate some of the fun and enthusiasm that children have for learning. But remember, even though we may be silly and have fun, I am still very serious about the goals of our course. You may want to read more about my teaching philosophy.
Computer Literacy: Every biologist should be familiar with word-processing (i.e., Word), database (i.e., Access), and spreadsheet (i.e., Excel) software. Informational Technology Services offers many interesting workshops that you should consider if you need to improve your computing skills.
Sequencing of the Arabidopsis genome has provided a number of insights into plant signal transduction and revealed interesting differences with other multicellular organisms. Whereas G-protein-coupled receptors (GPCRs) predominate in Drosophila, Caenorhabditis elegans, and humans, there are relatively few GPCRs in Arabidopsis (Arabidopsis Genome Initiative, 2000; Venter et al., 2001). Furthermore, multiple genes encode each of three heterotrimeric G-protein subunits (α, β, and γ) in animals, whereas Arabidopsis appears to have single genes encoding α and β subunits and two genes encoding γ subunits (Arabidopsis Genome Initiative, 2000; Mason and Botella, 2001; Venter et al., 2001). However, genes encoding Gγ have low sequence similarity, and the presence of additional Gγ genes in the genome cannot be excluded (Mason and Botella, 2000).
GPCRs interact with α, β, and γ heterotrimeric G-protein subunits (Hamm, 1998). During GPCR activation, the GPCR acts as a guanine nucleotide exchange factor, causing the α subunit to exchange guanosine-diphosphate (GDP) for guanosine-triphosphate (GTP). Subsequently, α-GTP separates from the βγ dimer, and disassociation of all three subunits from the receptor occurs. Both α-GTP and βγ transduce the signal of the activated receptor to downstream effectors. The GTP on the α subunit is hydrolyzed to GDP, inactivating the α subunit and allowing its reassociation with βγ to reform the inactive heterotrimer complex.
The definition of heterotrimeric G-protein subunits in plants is largely based on sequence similarities with animal heterotrimeric G-protein subunits. The Arabidopsis α subunit GPA1 has 36% identity and 73% similarity with animal α subunits (Ma et al., 1990), the β subunit has 50% identity with some animal counterparts (Weiss et al., 1994), and Arabidopsis γ subunits show some sequence similarity with human γ subunits (Mason and Botella, 2000, 2001). In addition to sequence homologies, biochemical similarities between animal and plant heterotrimeric G proteins have been demonstrated. The rice α subunit binds and hydrolyzes GTP (Seo et al., 1997). Furthermore, the α and β subunits are membrane localized (Weiss et al., 1997; Obrdlik et al., 2000), and the β and γ subunits have been shown to bind to each other in vitro (Mason and Botella, 2000, 2001). However, it has not been demonstrated that the plant α, β, and γ subunits can form a trimer, nor has any subunit been shown to bind to any particular putative GPCR in plants.
Previously, the functions of heterotrimeric G-protein signaling in plants have been largely inferred from pharmacological studies (Wu and Assmann, 1994; Jones et al., 1998; Ritchie and Gilroy, 2000). These studies have suggested functions for heterotrimeric G-protein signaling in the regulation of ion channels, gibberellin signal transduction, abscisic acid signaling, as well as other possible functions.
Recently, loss-of-function mutants in the heterotrimeric G-protein α subunits of rice and Arabidopsis have been described (Ashikari et al., 1999; Fujisawa et al., 1999; Ueguchi-Tanaka et al., 2000; Ullah et al., 2001; Wang et al., 2001). Although G-protein α-subunit null mutants from both species are completely viable, they show several developmental defects. The rice mutant exhibits shortened internodes, rounded seeds, and partial insensitivity to gibberellin, whereas the Arabidopsis gpa1 mutants have rounded leaves and altered sensitivity to a number of phytohormones, including gibberellin. Furthermore, gpa1 affects either cell division or cell elongation, depending on the organ type. Moreover, abscisic acid–regulated inhibition of stomatal opening requires GPA1 function.
Together with the Gα null alleles, plant Gα overexpression studies have documented the importance of G-protein signaling (Okamoto et al., 2001; Ullah et al., 2001). Ectopic overexpression of GPA1 increased cell division, led to formation of adventitious meristems, and increased developmental sensitivity to low levels of light. In contrast to Gα, neither gain-of-function nor loss-of-function mutants for any plant heterotrimeric G-protein β or γ subunits have been reported. Therefore, the relative physiological importance of these subunits has been unclear.
A genetic screen aimed at identifying genes functioning in the receptor-like kinase ERECTA (ER) signaling pathway was performed. The ER gene is predicted to encode a protein with 20 leucine-rich repeats in its extracellular domain, a single transmembrane domain, and an intracellular serine/threonine protein kinase domain (Torii et al., 1996). er mutants have pleiotropic phenotypes affecting the development of leaves, stems, flowers, and fruits (Rédei, 1962; Bowman, 1994; Torii et al., 1996). One of the mutants we identified in our screen encodes a mutant allele of the Arabidopsis heterotrimeric G-protein β subunit (AGB1). agb1-1 exhibits several defects, including short, blunt fruits, rounded leaves, and shortened floral buds. The phenotypic characterization of agb1-1 demonstrates that heterotrimeric G proteins play a role in plant development and contribute to our understanding of plant cell signaling.
Identification of agb1
We initiated a genetic screen to find mutants representing components of an ER signaling pathway, based on the hypothesis that other loci can be mutated to give an er phenotype. er mutants have rounded leaves, shortened petioles, pedicels, and stems, with flowers clustered together at the inflorescence apex (Rédei, 1962; Bowman, 1994; Torii et al., 1996). In addition, the siliques of er mutants are shorter, wider, and have a blunt tip. To test our hypothesis, we screened ethyl methanesulfonic acid–mutagenized populations of Arabidopsis for the clustered flower phenotype, which we followed with observations of the silique morphology. As summarized in Table 1, the mutants were placed into five complementation groups and named elk1 through elk5 (for erecta-like). Each elk mutant was mapped, as summarized in Table 2. Because of a similar map position and phenotypes, complementation testing was performed between elk1 and transport inhibitor resistant3 (tir3) (Ruegger et al., 1997), which revealed that they are allelic. Therefore, elk1 alleles were renamed tir3-101 to tir3-110. The remainder of the elk mutants do not map near previously described mutants with similar phenotypes. As a first step toward ascertaining which mutants represent genes that may function with ER, we performed silique length measurements of elk mutants (Table 3). All of the elk loci, except elk4, have a shorter silique than does a complete loss-of-function er reference allele. Therefore, tir3, elk2, elk3, and elk5 mutants were not studied further because they likely function in either separate and/or additional developmental pathways, with ER controlling silique shape. However, elk4, represented by a single allele, has a silique length almost identical to that of er. Therefore, we performed more detailed studies with elk4. Upon molecular cloning of the ELK4 locus, elk4 was renamed agb1.
Summary of elk Mutants
Map Positions of elk Mutants
Analysis of elk Mutant Silique Lengths
agb1 Affects Silique Morphology
A striking phenotype of agb1 is its silique morphology. Figure 1 shows that the development of the silique is altered in agb1 and is similar in appearance to the silique morphology of er. The tip of the wild-type silique has an acute appearance, because the width of the valves tapers apically and the valves contact the style at its base. Moreover, the style begins near the distal end of the valves and projects well past them. In contrast, the silique tips of agb1 and er plants have a blunt appearance. The valves do not taper apically and contact the style at its midpoint, and styles do not extend as far past the valves as they do in the wild type.
agb1 and er Cause Similar Changes in Silique Morphology.
(A) Excised siliques from wild-type plants (WT; ecotype Columbia [Col]), agb1-1, er-105, and agb1-1 er-105 double mutant. Bar = 5 mm.
(B) Scanning electron micrographs of silique tips from wild-type plants (WT; Col ecotype), agb1-1, er-105, and agb1-1 er-105 double mutant. stg, stigma; sty, style; v, valve. Bars = 400 μm.
The similarity in the silique morphology between agb1 and er prompted us to analyze the morphology of the double mutant silique. In Figure 2 and Table 4, statistical analysis of quantitative measurements of silique length, silique width, pedicel length, and silique tip angle is presented. In the single agb1 or er mutants, the length of the silique is decreased, whereas the width is increased. Furthermore, the angle of the fruit tip is smaller in agb1 and er than it is in the wild type, reflecting the blunt tip morphology. However, agb1 pedicel length is slightly longer than it is in the wild type, whereas er has a shorter pedicel. Double mutants were constructed to examine the interaction between agb1 and er in the control of silique form. Of the silique and pedicel traits measured, the silique length of the double mutant was significantly different from that of er, whereas the pedicel length, silique width, and angle of the silique tip were not significantly different from those of the er single mutant.
Morphometric Analysis of agb1 and er Silique Traits.
Comparison of silique and pedicel traits among wild-type plants (WT; Col ecotype), agb1-1, er-105, and agb1-1 er-105 double mutant. The first five fruits from eight to 10 plants were measured for each genotype. Means ±sd are in millimeters and were for 40 to 50 siliques per genotype. *Significantly different from the wild type (P ≤ 0.001). **Significantly different from the wild type (P ≤ 0.00001, Student's t test). er-105 is not significantly different from the double mutant for pedicel length (P ≥ 0.057, Student's t test) or silique width (P ≥ 0.301, Student's t test), but it is significantly different from the double mutant for silique length (P ≤ 0.00001, Student's t test). er-105 is significantly different from agb1-1 for pedicel length (P ≤ 0.00001, Student's t test) and for silique width (P ≥ 0.034), but it is not significantly different from agb1-1 for silique length (P ≥ 0.642, Student's t test). agb1-1 is significantly different from the double mutant for pedicel length (P ≤ 0.00001, Student's t test), silique length (P ≤ 0.00001, Student's t test), and silique width (P ≤ 0.00001, Student's t test).
In addition to the alterations in silique morphology, agb1 causes a modest shortening of floral bud length, as shown in Figure 3 . The floral buds at the inflorescence apex appear more tightly clustered together than they are in the wild type, but not as clustered as in er. This is because agb1 does not greatly affect pedicel or stem length, which are both shortened in er mutants (Bowman, 1994; Torii et al., 1996). Stem length is slightly decreased in agb1 relative to the wild type, but it is not as short as in er, as shown in Figure 3B and Table 5. The agb1 er double mutant is shorter than either agb1 or er single mutants.
agb1 Affects Flower Shape and Inflorescence Length.
(A) Comparison of inflorescence apices among wild-type plants (WT; Col ecotype), agb1-1, er-105, and agb1-1 er-105 double mutants ∼5 days after flowering. Bar = 1 mm.
(B) Comparison of 5-week-old adult plants. Bar = 2 cm.
agb1 plants also have rounded leaves and short petioles, which is shown by a comparison of rosette leaves in Figure 4 . From our initial comparisons of leaf characteristics between agb1 and the wild type, the ninth leaf appeared to show the greatest differences between the two genotypes. Therefore, we chose the ninth leaf for performing quantitative measurements. The statistical analyses of petiole length, lamina length, and lamina width are presented in Table 6. agb1 has petioles ∼50% as long as those in the wild type, whereas the lamina in agb1 is ∼125% wider and ∼85% as long as that in the wild type. These traits contribute to giving the agb1 rosette a more compact, rounded appearance than that of wild-type plants (data not shown).
agb1 Affects Leaf Shape.
Comparison of excised leaves from 4-week-old wild-type plants (WT; Col ecotype), agb1-1, er-105, and agb1-1 er-105 double mutants. The first leaves through the 10th leaves are seen left to right. Bar = 1 cm.
Because agb1 and er affect both leaf shape and petiole length (Figure 4, Table 6), we addressed the interaction between agb1 and er in the control of leaf shape by analyzing the agb1 er double mutant. With one exception, the double mutant has a shorter petiole, a shorter lamina, and a wider lamina than does either single mutant. The exception is that leaf width in agb1 and the double mutant do not show a statistically significant difference.
Measurements of Silique Tip Angles
Analysis of agb1 Inflorescence Length
Analysis of agb1 Leaf Traitsa
agb1 Encodes a Mutant Heterotrimeric G-Protein β Subunit
Molecular identification of the agb1 mutant was performed by following a positional cloning strategy, as illustrated in Figure 5 . agb1 was found to map between the markers F3L17(4800) and nga1107 on the bottom arm of chromosome four. F2 plants that were recombinant for these flanking markers were assayed with additional markers internal to the first pair to define a smaller pool of recombinants. This process was repeated in an iterative manner until the genomic region encompassing the AGB1 locus was delimited to 61 kb. At this point there was one recombinant on the centromeric side of AGB1 and two recombinants at a marker telomeric to AGB1.
Mapping of the AGB1 Locus to a 61-kb Interval.
Above the chromosome are the genetic markers used in the mapping studies. Below each marker are the number of recombinants observed for that marker out of the total number of chromosomes analyzed. The 61-kb mapping interval contains 16 genes. The transcriptional orientation and relative size of each gene are shown by arrows.
Sixteen annotated transcription units were present in the 61-kb interval. We transformed agb1-1 with each of the 16 genes to test for complementation. The genes were tested singly, or in three cases, two adjacent genes were tested simultaneously. On the basis of the gene annotation, polymerase chain reaction (PCR) products amplified with the wild-type Columbia (Col) ecotype genomic DNA template were cloned into a binary vector for Agrobacterium-mediated transformation. We found that a genomic fragment containing only a gene encoding a heterotrimeric G-protein β subunit was able to complement agb1-1, as documented in Figure 6A . The cDNA for this gene was previously identified and named AGB1, for Arabidopsis Gβ 1 (Weiss et al., 1994). Thirty independent transformants were obtained with the AGB1-containing construct, all of which showed complementation. For example, the rounded leaf phenotype of agb1-1 was restored to its normal shape. Also, the blunt silique tip of agb1-1 was complemented, having the acute tip seen in wild-type fruits. DNA gel blot analysis confirmed the presence of the construct used to complement agb1 in the genome of complemented plants (data not shown).
Complementation of agb1 and Identification of the Molecular Lesion in agb1-1.
(A) Comparison of phenotypic traits among wild-type plants (WT; Col ecotype), agb1-1, and complemented plants (homozygous agb1-1 transformed with pCAMBIA2300-T4L20.4). At top, rosette phenotypes of 3-week-old plants. Bar = 1 cm. At bottom, silique tip phenotypes from each genotype. Bar = 1 mm.
(B) At top, the AGB1 locus is comprised of six exons and five introns. Sequence analysis of agb1-1 genomic DNA and AGB1 cDNA from agb1-1 showed that the molecular lesion is a transition mutation altering the splicing donor site at the 5′ end of the first intron of AGB1. At bottom is the mature AGB1 mRNA from agb1-1.
(C) A portion of the AGB1 genomic DNA sequence flanking the first exon/intron junction, showing the molecular lesion in agb1-1. Above the exon DNA sequence, the amino acid residues for the last four entire codons before the first intron are shown.
agb1-1 Is a Null Allele
AGB1 genomic sequence in agb1-1 was analyzed, and a transition mutation at the splicing donor site of the first intron was found, as shown in Figure 6B. Sequencing of AGB1 cDNA isolated from agb1-1 revealed that the mutation results in a failure to splice out the first intron. RNA gel blot analysis of steady state levels of AGB1 in agb1-1 showed that the transcript is slightly larger in size than it is in the wild-type controls. The size of the mRNA from agb1-1 plants corresponds to the size of the transcript if the first intron is not removed. Failure to splice out the first intron appears to destabilize the transcript, because AGB1 mRNA is less abundant in agb1-1 (Figure 7) . When conceptually translated, the mature mRNA in agb1-1 is predicted to express the first exon, which consists of 35 amino acids, and then undergo a frameshift resulting in the addition of 20 novel amino acids and a premature stop. The predicted AGB1 protein in agb1-1 is truncated before the first of the seven WD-40 repeats that comprise AGB1 (Weiss et al., 1994). Thus, agb1-1 is likely to be a null allele.
AGB1 mRNA Is Larger in Size and Less Abundant in the agb1-1 Background.
RNA gel blot analysis of steady state levels of mRNA expression in wild-type plants (WT; Col ecotype), er-105, and agb1-1. The blot was sequentially hybridized with AGB1 and ER32P-labeled probes. At top, AGB1 steady state mRNA levels. The arrow indicates the wild-type AGB1 mRNA; the arrow with the asterisk indicates the aberrantly spliced AGB1 mRNA seen in agb1-1. At center, ER steady state mRNA levels. At bottom, ethidium bromide (EtBr)–staining of RNA to show loading. When normalized for loading, AGB1 transcript in er-105 is 90% of that in the wild type, AGB1 transcript in agb1-1 is 11% of that in the wild type, ER transcript in er-105 is 0% of that in the wild type, and ER transcript in agb1-1 is 74% of that in the wild type.
Expression of ELK4 (AGB1) was found in all tissues tested (Figure 8) . agb1-1 has the shortened, blunt silique phenotype of er but not the shortened pedicel phenotype. Consistent with the phenotypes observed in agb1-1, expression was high in the silique relative to the pedicel.
AGB1 Is Expressed in All Tissues Examined and Is Highest in Siliques.
At top, AGB1 mRNA signals from RNA gel blot analysis of wild-type plant tissues with a 32P-labeled AGB1 probe. From left, roots of 3-week-old plants, rosette leaves of 3-week-old plants, stems of 4-week-old plants (2 to 5 cm from the apex), unopened flower buds, open flowers (including pedicels) from 4-week-old plants, pedicels from stage 15/16 fruits (stages according to Smyth et al. ), and stage 15/16 siliques. At bottom, ethidium bromide (EtBr)–staining of RNA to show loading.
agb1 affects the shape of leaves, floral buds, and fruits. The silique phenotypes of agb1-1 are reminiscent of those exhibited by the er mutant (Rédei, 1962; Bowman, 1994; Torii et al., 1996), which raises the possibility that AGB1 functions in a common silique developmental pathway with ER. This possibility was addressed by examination of agb1 er double mutants. The two simplest cases are that either AGB1 and ER function in parallel pathways or they function in a common developmental pathway. For example, if AGB1 and ER function in parallel pathways, the double mutant would be expected to show a more severe phenotype, evidenced by shorter, wider siliques. For this test to be meaningful, Arabidopsis fruits must have the capacity of being either shorter or wider than those seen in agb1 or er mutants. This requirement is satisfied because mutants have been reported that make the silique either shorter or wider than either agb1 or er. For example, tir3 mutants have siliques that are <60% as long as those in the wild type (Ruegger et al., 1997), and overexpression of CYP78A9 results in Arabidopsis fruits that are 40% wider than are er fruits (Ito and Meyerowitz, 2000).
Statistical analysis of the agb1 er double mutant shows that the silique is significantly shorter than that in either the agb1 or er single mutant. Similarly, for leaf and stem phenotypes, the double mutants were more severe than were the er or agb1 single mutants. These observations suggest that AGB1 and ER function in parallel pathways that control the development of these traits. If they do function in parallel pathways, it is possible that agb1 simply phenocopies some aspects of er mutants but that each mutant has a different physiological basis for the similar aspects of their morphology.
However, statistical analysis of silique width, as well as tip angle measurements of agb1 er double mutants, showed these traits are not significantly different from those of either agb1 or er single mutant siliques. Futhermore, pedicel length in the double mutant is not significantly different from that in the er single mutant. These observations support the hypothesis that AGB1 functions in a common developmental pathway, with ER controlling these characteristics.
If AGB1 and ER function in a common developmental pathway, they might function in a common signal transduction cascade. Many signal transduction pathways modulate the expression of genes to bring about a cellular response (Wodarz and Nusse, 1998). One possibility we considered was that ER is transcriptionally activated by a GPCR pathway involving AGB1. If this case were true, we would expect to observe lower levels of ER expression in agb1-1. However, steady state levels of ER mRNA are similar in an agb1-1 background. A second possibility is that AGB1 expression requires functional ER. In this case, we would expect to see loss of AGB1 expression in an er background. This scenario is unlikely because steady state levels of AGB1 mRNA are only slightly reduced in er.