Plant Molecular Biology 39: 577–592, 1999.
© 1999 Kluwer Academic Publishers. Printed in the Netherlands.
Differential expression of genes encoding the light-dependent and
light-independent enzymes for protochlorophyllide reduction during
development in loblolly pine
Jeffrey S. Skinner
and Michael P. Timko
Department of Biology, University of Virginia, Charlottesville, VA 22903, USA (
author for correspondence);
Current address: Department of Forest Science, Oregon State University, Corvallis, OR 97331-7501, USA
Received 26 May 1998; accepted in revised form 29 September 1998
Key words: chlorophyll, gymnosperm, loblolly pine, NADPH:protochlorophyllide oxidoreductase, protochloro-
The expression patterns of the two distinct subfamilies of genes (designated porA and porB) encoding the light-
dependent NADPH:protochlorophyllide oxidoreductases (PORs) in loblolly pine (Pinus taeda L.) were examined.
Transcripts arising from both gene subfamilies were shown to be present at high levels in the cotyledons of dark-
grown pine seedlings and to a lesser extent in their stems. Exposure of dark-grown seedlings to light resulted
in increased levels of both porA and porB transcripts, as well as increased levels of mRNAs encoding other
photosynthesis-related gene products, suggesting that they are under a common mode of regulation. Relative levels
of the porA and porB transcripts were similar in seedling cotyledons and primary needles of two-month-old pine
trees, whereas only porB transcripts were present at a signiﬁcant level in mature secondary needles of two-year-
old trees. Immunoblot analysis showed that the 37 kDa PORA protein was most abundant in dark-grown tissues,
decreased dramatically upon exposure to light, but could still be detected at low levels in light-grown seedlings.
In comparison, levels of the 38 kDa PORB protein were not signiﬁcantly changed upon transfer of dark-grown
tissues to light. While both PORA and PORB were detected in cotyledons and primary needles, only PORB could
be detected in mature needles. Transcripts derived from the three plastid genes, chlL, chlN,andchlB, encoding
subunits of the light-independent protochlorophyllide reductase were detected in the cotyledons and stems of dark-
grown seedlings, and in mature needles. The highest levels of chlL, chlN,andchlB transcripts were detected within
the top one-third of the stem and decreased gradually towards the stem/root transition zone. Correspondingly, the
highest levels of light-independentchlorophyllformation took place near the top of the hypocotyl. A similar pattern
of expression was observed for other photosynthesis-related gene products, including porA and porB. Our results
suggest that many aspects of the light-dependent, tissue-speciﬁc and developmental regulation of POR expression
ﬁrst described in angiospermswere already established in the less evolutionarilyadvanced gymnosperms. However,
unlike angiosperms, light is not the dominant regulatory factor controlling porA expression in these species.
Abbreviations: nt, nucleotide(s); PLB, prolamellar body; POR, light-dependent NADPH:protochlorophyllide
oxidoreductase; UTR, untranslated region; chlide, chlorophyllide; pchlide, protochlorophyllide
Our understanding of how chlorophyll biosynthesis is
regulated in vascular plants and the role its formation
and accumulation play in the control of chloroplast
development and overall plant photomorphogenesis
has advanced considerably in recent years [11, 33,
34]. It is now generally accepted that chlorophyll
formation is regulated, at least in part, by the reduc-
tion of protochlorophyllide (pchlide) to chlorophyllide
(chlide). During evolution, two distinct biochemi-
cal routes for pchlide reduction evolved in nature.