Recent Results


















 

 

 

 

 

 

  

 

 

  

  

 

 

 

 

 

 

 

 

 

 

 

 

 

 

The bulk of the stuff from the sections below until now (late 2015) are in the publication links:

We are in the beginning stages of developing a greenhouse based system to analyze the response of sunflowers to weeds- specifically, two canola varieties and two amaranth species (GR-Palmer amaranth, and a grain amaranth (cv Plainsman). So far, it looks like the canola has a greater impact on the sunflower than either of the amaranths, but - at least by the harvest date (2 months after planting) we can see significant differences in leaf area, stem diameter and leaf number for all treatments relative to control at least for the population of sunflower derived from US-grown plants. There does appear to be some epigentic responses involved since the seeds collected from the winter nursery in Chile have a lot more variability in phenology and their response to weeds seems to be muted relative to the US-grown plant seeds. 





We recently completed a study where we used microarray analysis to identify physiological processes affected by weed competition in corn. These initial studies were done on the top-most leaf of corn at the V11-14 stage of growth (well after the corn had overtopped the velvetleaf). Microarray analysis identified numerous genes involved in photosynthesis, protein degradation, carbon usage, cell division, and ethylene and auxin signaling that were depressed by weed competition.  The observation that photosynthesis was depressed was surprising since the leaf material collected should not have been experiencing any competition for light. Thus, either the presence of weeds in the sub-canopy can have a detrimental effect on whole plant photosynthesis (possibly by limiting crucial nutrients?), or the presence of the weed permanently altered these systems in the crop. These studies have been accepted for publication in Weed Science. Further studies are planned to determine how weeds effect corn during earlier stages of growth when they are more likely to be directly competing for light. Studies are also under way to determine what long term effects weed competition has on corn by observing the changes in the transcriptome when the weed is removed following the critical period during which weeds are known to have a permanment effect on crop yield. Additionally, a comparison of the response to shading and high density verses control density was completed. Changes in PEP carboxylase expression was different between the two stresses.

 

We have also used 24,000 element cotton microarrays developed by CSIRO to study the effect of crop competition on the velvetleaf. Both cotton and velvetleaf are members of the Mallow family and it was expected that many of the cDNAs on the array would hybridize to the labeled velevtleaf transcripts. Unfortunately only about a 1/3 of the probes hybridized at levels at least a standard deviation above background. However, that still meant that the expression of over 5,000 characterized velvetleaf genes could be studied. The results also demonstrated that genes involved in ethylene signaling, cell division, and photosynthesis were preferentially expressed in velvetleaf competing with corn.  

In our quest to understand how flowering signaling impacts fall dormancy, we have recently cloned cDNAs for FT and a related FT-LIKE gene from leafy spurge. The FT-LIKE gene appears to be constitutively expressed, whereas the FT orthologue appears to be diurnally regulated (see F (flowering) D (leaves collected at 11:00 AM) and N (leaves collected at 12:00 PM- 3 hr after sundown). We have correlated the loss reduced expression of FT with onset of fall dormancy as was demonstrated in poplar. We have also collected spurge plants from different latitudes to see if there are threshold daylength differences for when FT expression is lost. Interestingly, our greenhouse plants that have been vegetatively propogated for many years and have lost the ability to flower show a constant level of FT expression (NF on the figure to the left) whereas FT is diurnally regulated in our outdoor grown plants which do flower.

In a related experiment, we have cloned both cDNA and genomic copies of a MAF3/1 orthologue from spurge and are in the process of looking to see if there are similar MAF-like genes clustered together in the genome of leafy spurge. Interestingly, there appears to be numerous copies of this gene with some probable differential splicing that may be differentially regulated in a sasonal manner.

The microarray data is now in for the seasonal bud dormancy!  A text file of the initial analysis can be down loaded below (click on the hypertext words "Microarray"). Several interesting obnservations were made when we compared our results to two other seasonal microarrays from poplar and raspberry.  One was there is a common MADS Box transcription factor that is differentially regulated during dormancy transitions. A similar series of MADS-box gene (now called DORMANCY ASSOCIATED MADS_BOX or DAM) was found to be deleated ina mutant variety of peach that does go dormant in the fall. The EST for this gene in spurge (and cassava) is truncated to the 1st exon plus an additional 8 ammino acids that does not appear to be the result of read-through into the first exon. We have cloned this gene and several other variants and found them to be differentially regulated through dormancy transitions.  Interestingly, there appears to be several introns within the 5'UTR of the gene (click here for details on sequences and expression of DAM variants). We have prepared a construct to over and underexpress this gene in arabidopsis, poplar, and leafy spurge. Preliminary results appear to indicate that the short DAM1a gene causes late flowering when expressed in arabidopsis. We are also collaborating with Dr. Sibum Sung (formerly from Rick Amasino's lab) to use chromatin immunoprecipitation (ChIP) to look for chromatin modification associated with dormancy transitions and to see if DAM binds to regulatory sequences in FT.

Vijay has recently identified transgenic arabidopsis plants that were transformed with a construct of GUS driven by the leafy spurge STM promoter. STM is a key gene involved in meristem development and is generally only expressed in growing meristems. The key difference between the leafy spurge STM promoter and the arabidopsis STM promoter is the presence of a tuber-sprecific sugar-responsive element in the leafy spurge version. It should be noted that leafy spurge forms buds on it's roots whereas arabidopsis does not. Consequently we were pleased to see GUS expression in the meristematic region of the trangenics as well as GUS expression in the root pericycle cells. A construct that knocks out the tuber-sprecific sugar-responsive element has been prepared and  transformed into arabidopsis.  Interestingly, it does not appear to affect expression in the roots of the transgenic plants. Another interesting observation from this work is that there are several variants of STM in leafy spurge. One variant (STM4) is expressed in young and old roots (including roots with no visable buds), hypocotyl, and meristem, and the other (STM1) is only expressed in the young roots and hypocotyl. We are currently collaborating with Dr. Jan's lab to confirm expression of STM in roots of leafy spurge by in situ hybridization.

Microarray results:  Note, differential expression is shown as the average normalized log2 expression from all samples listed as paradormant (all August and September samples plus two October samples), endodormant (the remainder of the October samples plus three November samples) and ecodormant (the remainder of the November samples and the December samples). We have also blasted our leafy spurge contigs against the sequences from poplar, raspberry, potato, and grape microarrays that heve been used in other dormancy studies to ID common differentially expressed genes in these perennial species. A table showing the various differentially-expression genes and their relative trends through dormancy transitions can be downloaded by clicking here. STable 1, STable 2, STable3a, STable3b, STable3c, STable4, STable5, STable6, STable8, SFigures.

We have completed our study on the function of the conserved hairpin sequence adjacent to the ATG in the 5' UTR of CYCLIN D3-2 gene. This sequence appears to target cleavage of the transcript in a manner dependent on DICER. Interestingly, it appears that the three base pairs in the loop section are required for cleavage and not the hairpin itself. We are also checking to see if cytokinin (which induces CYCLIN D3-2) causes any change in the accumulation of the cleaved fragment.

The recent visit to my lab by Dr. Maria Santana has resulted in a series of excellent microarray studies on the progression of Xam infection in cassava and leafy spurge.  The leafy spurge data can be down loaded here.  DATA