trans

DEVELOPMENTAL VARIATION AND CARBOHYDRATE STORAGE 167
FIGURE 7.9 Electron micrographs of Toxoplasma gondii. (A) T. gondii tachyzoite, showing a nucleus (N) and various
organelles of the ‘apical complex’: C, conoid; R, rhoptries; M, micronemes. Dense granules (DG) are involved
in constitutive secretion, releasing parasite proteins into the parasitophorous vacuole. (B) The bradyzoite stage
displays a similar complement of organelles (albeit organized in a slightly different manner), but is characterized
by the appearance of amylopectin/floridean starch granules (PG). (C) Dozens of T. gondii enclosed within a single
tissue cyst. Note the prevalence of starch granules within individual bradyzoites, and the proteoglycan wall
surrounding the bradyzoite cyst. Electron micrographs provided courtesy of David J. Ferguson, Oxford University.
The first committed step in the mannitol
pathway is carried out by mannitol 1-phosphate
dehydrogenase (M1PDH), which converts fructose
6-phosphate into mannitol 1-phosphate.
M1PDH is developmentally regulated by a
homodimeric protein, which inhibits activity.
Removal of this homodimer by denaturation,
pH shift, or addition of a polyclonal antibody
raised against the inhibitory protein restores
M1PDH activity. A mannitol 1-phosphatase
irreversibly converts mannitol 1-phosphate to
mannitol and completes half of the cycle. This
phosphatase is highly specific for its substrate,
and has also been detected in Toxoplasma
gondii and Cryptosporidium oocysts.
Temperature acts as an interesting regulator
of mannitol production. Both M1PDH and
the phosphatase are highly active at 41°C in
E. tenella (the typical body temperature of the
avian host), but essentially inactive at 25°C
(outside of the host). In contrast, enzymes
involved in mannitol catabolism – including
mannitol dehydrogenase (MDH) and hexokinase
(the same enzyme initiating the glycolytic
pathway) – are substantially more
active at ambient temperatures, correlating
with the degradation of mannitol upon oocyst
sporulation.
Specific inhibition of M1PDH by a 3-
nitrophenyl disulphide (nitrophenide) leads
to defective sporulation, validating the pathway
as essential for completion of the parasite
life cycle. Despite the formidable energy
storage capabilites of starch granules, the
BIOCHEMISTRY AND CELL BIOLOGY: PROTOZOA