Proteolytic enzyme activities in viviparous-1 and normal kernel tissues during development

--Kien Tjhen, J.D. Smith and D. G. Bai

Electron micrographs reveal that protein bodies in aleurone tissue, but not endosperm, are seriously degraded by 25 days after pollination (DAP) in viviparous kernels. As expected, these protein bodies appear to be intact through maturity in normal kernels.

Enzymatic activities of carboxypeptidase (CP) and endopeptidase (EP) were determined from purified extracts of aleurone, endosperm, and embryo tissue of developing maize kernels at 15, 20, 25 and 35 DAP. These ages correspond to stage 4, stage 6, mature embryos and black layer, respectively (MNL 61:40). The activities of CP and EP were investigated using routine TCA methods, at pH 5.0 with phenylalanine and at pH 5.4 with zein as respective substrates, followed by ninhydrin reaction for CP.

The CP and EP in vp/vp and normal kernels showed similar patterns of activities in all tissues. The activity of CP was about 0.06 mol/g/h and EP was about 0.1 mol/g/h during early seed development (15 DAP). The highest activity was seen in embryo tissue for CP (0.085 mol/g/h) and in aleurone tissue for EP (0.6 mol/g/h) at 25 DAP.

The activity of CP in aleurone tissue decreased significantly after 15 DAP, indicating that this enzyme was very active in aleurone tissue during early development. The activities of EP in aleurone and embryo tissues showed that this enzyme was very active at this point but turned off after 25 DAP. The endosperm tissue had low or almost negligible CP and EP activities at all stages of seed development.

Table 1. Carboxypeptidase (CP) activity in viviparous-1 and normal maize kernel tissues.
CP activity (mol/g/h)
  DAP Embryo Endosperm Aleurone
  15 0.0588 0.0671 0.0632
Mutant 20 0.0646 0.0579 0.0605
  25 0.0960 0.0201 0.0215
  35 0.0379 0.0028 0.0037
  15 0.0729 0.0926 0.0677
Normal 20 0.0696 0.0562 0.0662
  25 0.0824 0.0229 0.0242
  35 0.0525 0.0040 0.0219

Table 2. Endopeptidase (EP) activity in viviparous-1 and normal maize kernel tissues.
CP activity (mol/g/h)
  DAP Embryo Endosperm Aleurone
  15 0.0983 0.1829 0.1428
Mutant 20 0.2039 0.1038 0.1629
  25 0.6525 0.1234 0.9086
  35 0.6289 0.0820 0.4062
  15 0.0690 0.1785 0.1149
Normal 20 0.3142 0.2298 0.3298
  25 0.5136 0.1462 0.9129
  35 0.5040 0.0641 0.5645

Although some differences were observed, the similarities in the activity patterns of these proteolytic enzymes indicate that the degradation of aleurone protein bodies in vp/vp kernels is not due to Vp regulation of these enzymes.

Please Note: Notes submitted to the Maize Genetics Cooperation Newsletter may be cited only with consent of the authors

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