Department of Botany & Plant Sciences

Harley M.S. Smith

Assistant Professor and Assistant Plant Cell Biologist (Ph.D., 1998, Michigan State University)
Office: 4202B Genomics Building
Phone: (951) 827-2643
Fax: (951) 827-5155
Email: harleys@ucr.edu

Areas of Expertise

• Cell and Developmental Biology
• Genetics

Background
Regulation of Inflorescene Architecture in Arabidopsis
Selected Publications (Bibliography page)

​Background

My college education began at Cabrillo Community College in Aptos, CA (www.cabrillo.edu) after leaving the construction business to pursue my interests in biology. I transferred to the University of California, San Diego (www.ucsd.edu), where I received a B.S. in Biochemistry and Cellular Biology. I received a Ph.D. in Genetics at Michigan State University in the Department of Energy-Plant Research Laboratory (www.prl.msu.edu). I worked on my thesis project in Dr. Natasha Raikhel’s Laboratory (www.cepceb.ucr.edu/members/raikhel.htm) where I studied the function of a nuclear localization signal receptor, AT-IMPORTIN ALPHA.

After receiving my Ph.D., I pursued post-doctoral studies in plant development in Dr. Sarah Hake’s Laboratory (www.pgec.usda.gov/Hake/SHresearch1.html) at the University of California, Berkeley-USDA Plant Gene Expression Center (www.pgec.usda.gov). My background in cell biology allowed me to bring new approaches to basic problems in plant developmental biology that complemented the genetic approaches used in Dr. Hake’s laboratory. I was awarded a three year Post-doctoral Fellowship from the National Institute of Health (NIH) that involved developing a project to gain insight into the function of the KNOTTED1-like homeobox (KNOX) transcription factors in maize and Arabidopsis. During the first phase of my project, we showed that KNOX proteins form complexes with the BEL1-like ( BELL) family of homeodomain proteins. Studies from our laboratory at the CEPCEB indicate that inflorescence development requires the activities of specific KNOX-BELL heterodimers during reproductive development in Arabidopsis.

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​Project 1: Marker assisted breeding in avocado

The objective of this project is to accelerate the avocado-breeding program by screening for favorable reproductive phenotypes at the seedling stage.  In this project, we are implementing a marker assisted selection (MAS) scheme to identify trees that produce fruit containing higher nutritive values.  Currently, we are screening 1600 F2 seedlings derived from 8 F1 individuals that produce fruit with high nutritive value.  Our goal is to identify approximately 50 individual seedlings that contain more than one nutritive marker.  These seedlings will then be incorporated into the avocado-breeding program headed by Dr. Mary Lu Arpaia.  Once the trees reach reproductive maturity, analyze once the fruits for nutritive value as well as fruit quality.  To accomplish this project, I received a grant from the California Avocado Commission titled "Implementing marker-assisted selection for biochemical phenotypes in avocado".

Figure 1: DNA concentration for each sample was measured using a Nanodrop instrument and successively diluted to 1-20 ng for the optimal template concentration for TaqMan SNP genotyping assay. This assay was performed with the putative lutein (LUT5) gene that had been previously cloned in the F1 mapping population. The PCR conditions were optimized on a BIO-RAD CFX96 real-time PCR instrument at the UCR core genomics facility. Results show that three control genotypes (control 1 = homozygous for allele a; heterozygous (ab), and control 2 = homozygous for allele b) can be successfully discriminated using this assay.

Project 2: Variety improvement in avocado

Another project is to utilize an avocado F1 population and selected germplasm genotypes to map horticulturally favorable traits in avocado. This is a collaborative project with CE-Specialist-Dr. Mary Lu Arpaia in the Dept. of Botany and Plant Science at UCR. The overall goal of this research is develop new improved avocado varieties that meet the needs of California avocado growers.

Figure 2: Fruit load promotes dormancy of the shoot apex during vegetative growth. (A) The apex of shoots without fruit actively produced leaves.   (B) Close up image of a shoot tip with fruit. The white arrow points to the main shoot apex, which appeared to have entered a dormant state. Also, the axillary buds are typically smaller and flattened in (B) shoots bearing fruits, compared to (A) shoots without fruits.

My laboratory is currently focused on using the F1 mapping population to identify traits associated with alternate or biennial bearing. Results from my lab show that the fruit inhibit growth the shoot (Figure 2). As a result, shoots bearing fruits produce fewer reproductive nodes, which likely contributes to the decline in flowering the following year. To understand the developmental basis of alternate bearing, my research program utilizes a multi-pronged approach, which integrates molecular, cellular, and genomic approaches to understand how fruit/crop load alters the growth and development of the shoot/tree. Preliminary results from my laboratory indicate that the avocado F1 mapping population displays marked variation in phenotypes associated with alternate bearing (Figure 3 and 4). In addition, this population displays variation in phenotypes attributed drought tolerance, tree architecture and fruit size, shape and quality. In collaboration with Dr. David Kuhn at the USDA in Miami, FL, we are utilizing an avocado Infinium SNP array to map the alternate bearing traits as well as other favorable traits. The avocado SNP array contains 6000 SNPs spanning the avocado genome. The goal is to not only identify genes responsible for controlling alternate bearing, but to use the SNPs associated with these traits in a marker assisted selection scheme. Through collaboration with the International Avocado Working Group, we hope to create an avocado marker assisted tool kit for breeding optimal avocado varities that are superior to Hass.

Figure 3: Variation in shoot growth in response to fruit load in the F1-population.

(A-B) Genotype #3 (DSP#3), clone A. (A-B) Numerous shoots in the tree are initiating leaves. (C) However, not all shoots are growing. The red arrow points to the dormant bud in which shoot growth was completely inhibited by the fruit. (D-F) Genotype #127 (ASP1#127), clone A. (D-E) Image of tree shows numerous shoots are active. (F) Despite fruit load the shoots are relatively active. Red arrows mark the starting point of the spring and summer flush. Genotype #88 (ASP#88). Shoot bearing fruit produce vegetative nodes during the spring and summer flush. (H-I) In addition, 1-2 reproductive nodes are produced, which give rise to inflorescences.

Figure 4: Variation in the effect of fruit load on the reproductive potential of the shoot in the F1-population. Genotype #132 displayed a TAP. (A-B) Inflorescences fail to develop on shoots bearing fruit as well as (C) nearby shoots.

Genotype #63 displayed a SAP. (D-E) The production of inflorescences is inhibited on shoots bearing fruit. However, nearby (F) shoots produce numerous inflorescences. Red arrows point at shoots bearing fruit, while white arrows point at nearby shoots. F = Fruit.

Project 3: Regulation of shoot meristem function and fate in Arabidopsis

The objectives of this project is to determine the role of two related homeodomain transcription factors, PENNYWISE (PNY) and POUND-FOOLISH (PNF), in regulating the function and fate of the shoot apical meristem and inflorescence architecture in Arabidopsis. The pny pnf double mutant is unique in that these plants fail to initiate flowers and internodes in response to floral inductive cues. Genetic, molecular and genomic approaches were used to demonstrate that PNY and PNF regulate meristem maintenance by regulating boundaries in the shoot meristem. This regulatory event is crucial for maintaining stem cell identity in the meristem center and for establishing positional cues in the peripheral zone that is necessary for flower formation. My laboratory also showed that the function of the florigenic signal, FLOWERING LOCUS T (FT), is dependent upon PNY/PNF, suggesting that PNY/PNF integrate FT activity to mediate the floral transition in the shoot meristem. Gene expression profiling using the ATH1 microarray showed that PNY/PNF regulate the SPL/microRNA156 module, which acts to control phase change and flower meristem identity. Currently, we are developing a Chromatin Immunoprecipitation-Sequencing (Next Generation Sequencing) approach to identify genes directly regulated by PNY/PNF in order to further develop the gene regulatory networks controlling meristem function and identity.

Figure 5: Role of PNY and PNF during flower specification

A model is shown which displays the photoperiod and endogenous flower specification pathways. FT-FD and SPL3/SPL4/SPL5 (collectively referred to as SPLs) act in parallel to induce SOC1 and FUL as well as flower meristem identity genes during inflorescence development. FT requires PNY and PNF for specifying flower meristem identity. In addition, PNY and PNF regulate SPL3, SPL4 and SPL5 by repressing miR156 during inflorescence development. At the same time, PNY and PNF function with SPL3, SPL4 and SPL5 during the specification of flower meristem identity. The dashed line between PNY/PNF and miR156 indicates that the nature of this regulation is not known.