Transient conz1 Induction of FT-like genes in Maize Protoplasts

Kristina Stemler
Marquette University
Mentor: Dr. Jane Dorweiler

Flowering time genes play a vital role in the transition plants undergo from vegetative to reproductive growth.  Day length, or photoperiod, has been postulated to impact the regulation of flowering time in multiple plant species though very little is known at the molecular level in maize.  CONSTANS (CO), the proposed floral regulator from Arabidopsis (Arabidopsis thaliana) has been shown to activate FLOWERING LOCUS T (FT), a floral initiator.  Comparatively, less is known about maize (Zea mays), an important crop.  Unlike Arabidopsis, which is a long day plant, favorably producing an inflorescence under long day conditions, maize is considered day neutral, flowering regardless of day length.  However, conz1 has been shown to act in response to photoperiod, which is consistent with its ortholog, CO, in Arabidopsis.  This observation suggests that CONZ1 protein should act in a smiliar manner on FT-like genes in maize.

Based on the model of Arabidopsis floral induction, we proposed to transform 35S::conz1 cDNA engineered in the pART 7 plasmid into maize mesophyll protoplasts to test in vivo whether conz1 overexpression activates transcription of FT-like genes.  We were also interested in testing the effects of dark versus blue light incubation conditions after transient transformation on the amount of FT-like gene activation because previous studies have shown that different wavelengths of light can either stabalize or assist in the degredation of CO protein in Arabidopsis.

Successful transient transformation of protoplasts was shown through an upregulation in conz1 expression.  Transformation was most efficient in samples containing dilute protoplasts and minimal plasmid DNA. 

Neither an upregulation nor expression of FT-like was evident despite attempting to obtain mRNA expression through the use of multiple primer pairs.  An absence of FT-like expression levels therefore did not give any data to draw conclusions on the effects of different incubation conditions post transformation.  Due to failure of FT-like expression, we tried to determine if conz1 overexpression resulted in upregulation of a different putative downstream gene target, SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1-like (soc1-like).  soc1 also failed to overexpress in transformed protoplasts.  It is possible that CONZ1 protein acts as a transcriptional repressor in maize, instead of as an activator, thereby following rice (Oryza sativa) as a model.
 
 
 
 

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