(a) Describe along with illustrated diagrams the structure and function of mitochondria and endoplasmic reticulum. 8+7=15 (b)(i)…

(a) Mitochondria and endoplasmic reticulum: structure and function

Mitochondria: structure

• Shape and number: rod-shaped or spherical, about 0.5–1 µm wide. Plant cells hold from a few hundred to several thousand, most in active tissues such as meristems and phloem companion cells.
• Outer membrane: smooth and freely permeable to small molecules because of porin channels.
• Intermembrane space: protons pumped by the electron transport chain accumulate here.
• Inner membrane: folded into cristae (often tubular in plants). It is impermeable to ions, rich in cardiolipin, and carries the electron transport complexes I–IV and ATP synthase (F₀F₁ particles).
• Matrix: contains the enzymes of the TCA cycle and fatty acid oxidation, circular mtDNA, 70S-type ribosomes and granules.
• Genome: plant mtDNA is unusually large (about 200–2,500 kb; about 367 kb in Arabidopsis), recombines into subgenomic circles, and uses RNA editing. Most mitochondrial proteins are nuclear-encoded and imported through the TOM/TIM complexes, so the organelle is semi-autonomous.
• Origin: mitochondria descend from an α-proteobacterial endosymbiont.

Mitochondria: functions

• Aerobic respiration: pyruvate is oxidised by pyruvate dehydrogenase and the TCA cycle. The NADH and FADH₂ formed pass electrons down the electron transport chain, and ATP synthase makes ATP by chemiosmosis (Mitchell). This supplies most ATP in non-green tissues and at night.

Plant-specific features:

• Rotenone-insensitive external NAD(P)H dehydrogenases.
• A cyanide-resistant alternative oxidase, which releases energy as heat; aroids such as Sauromatum and Symplocarpus use it to volatilise odours that attract pollinators.
• Glycine decarboxylase of photorespiration, which turns two glycine into serine and releases CO₂.
• Other roles: carbon skeletons for amino acids (2-oxoglutarate for nitrogen assimilation), Fe–S cluster and vitamin synthesis, Ca²⁺ buffering, and the control of programmed cell death. Mitochondrial genes also cause cytoplasmic male sterility, as described in (b).

Endoplasmic reticulum (ER): structure

The ER is a continuous network of membranes enclosing one lumen, continuous with the outer nuclear membrane. It extends into plasmodesmata as the desmotubule, linking neighbouring cells.

• Rough ER (RER): flattened cisternae studded with 80S ribosomes on the cytosolic face.
• Smooth ER (SER): branched tubules without ribosomes, abundant in cells that secrete oils, resins or waxes.

ER: functions

Synthesis of secretory, membrane and vacuolar proteins (RER):

• A signal peptide is recognised by the signal recognition particle.
• The ribosome docks on the ER and the protein is threaded into the lumen as it is made (co-translational import).
• There it is folded by chaperones (BiP, calnexin/calreticulin), disulfide bonds are formed (PDI), and N-linked glycosylation begins.
• Quality control: misfolded proteins are exported and degraded (ERAD), and overload triggers the unfolded protein response.
• Storage: prolamin protein bodies in maize and rice endosperm bud directly from the ER. Oil bodies (triacylglycerol with oleosin) of oilseeds form from ER membranes.
• Lipid synthesis (SER): phospholipids, triacylglycerols, and sterol and wax precursors.
• Detoxification: cytochrome P450 enzymes metabolise herbicides and other xenobiotics.
• Calcium store: the ER holds Ca²⁺ for signalling.
• Biotechnology: the retention signal KDEL/HDEL is added to recombinant proteins in plant "molecular farming" to keep them in the ER and increase yield.

(b)(i) Male sterility and cytoplasmic inheritance

Cytoplasmic inheritance is the transmission of traits by genes in mitochondria or plastids. In most angiosperms these organelles come only from the egg. Such traits therefore show a reciprocal difference (A ♀ × B ♂ is not the same as B ♀ × A ♂), follow the mother, and do not segregate in Mendelian ratios.

Cytoplasmic male sterility (CMS) is the classic plant example. Rhoades first described it in maize (1933).

• Proof of cytoplasmic inheritance: a male-sterile plant pollinated by a fertile line gives male-sterile progeny. This continues through repeated backcrosses, even after the nucleus has been almost entirely replaced by the fertile parent's genome. The factor therefore lies in the cytoplasm.
• Cause: rearrangements in the mitochondrial genome create chimeric open reading frames. Their protein products disturb mitochondrial function in the tapetum and developing pollen, which have a very high energy demand.

Examples:

• Maize T-cytoplasm (Texas): the gene T-urf13 encodes a pore-forming membrane protein. It also made T-cytoplasm maize susceptible to the toxin of Bipolaris maydis race T, causing the 1970 US southern corn leaf blight epidemic.
• Rice wild-abortive (WA) cytoplasm: WA352 interacts with the nuclear-encoded COX11 and triggers premature tapetal death.
• Sunflower PET1 (from Helianthus petiolaris; basis of nearly all sunflower hybrids).
• Ogura CMS in Brassica (orf138, from radish).
• Pearl millet A1 cytoplasm (Tift 23A).
• Nuclear–cytoplasmic interaction: fertility is restored by dominant nuclear **restorer-of-fertility (Rf)** genes. Many encode PPR proteins that bind and suppress the CMS transcripts. So a plant is male-sterile only with S cytoplasm and rf rf. This combination is used in the three-line hybrid system: A-line (S, rf rf), maintainer B-line (N, rf rf) and restorer R-line (Rf Rf).
• Genic male sterility, by contrast, is inherited in Mendelian fashion through the nucleus.

(b)(ii) Molecular basis of sex determination in plants

Most flowering plants are hermaphrodite. About 6% of species are dioecious and about 7% monoecious. Separate sexes evolved many times, so the mechanisms differ.

1. Sex chromosomes and sex-determining genes (dioecious species)

• Heteromorphic XY: Silene latifolia has a large Y carrying three kinds of functions: a gynoecium-suppressing factor, a stamen-promoting factor and male-fertility genes. Deletion mapping of the Y shows this. In Rumex acetosa, sex depends on the X : autosome ratio.
• Young, homomorphic XY: in papaya the male-specific region of the Y (MSY) is only a few megabases. A slightly different Yʰ gives hermaphrodites, and YY is lethal.

Two-gene model (a male-sterility mutation plus a female-suppressor, linked on the Y):

• Asparagus: SOFF (suppressor of female function) and TDF1 (tapetal development, needed for anthers) lie on the Y.
• Kiwifruit: the Y genes are Shy Girl, a cytokinin-response regulator that suppresses carpels, and Friendly Boy.
• **One-gene model (persimmon, Diospyros lotus):** the Y-linked gene OGI makes a small RNA that silences the autosomal feminising gene MeGI. Males lack MeGI activity and so develop as male.
• ZW systems occur in wild strawberry (Fragaria virginiana).

2. Hormones and genes in monoecious cucurbits

Ethylene promotes female flowers and gibberellins promote male flowers. Ethrel therefore raises femaleness, and silver nitrate or GA induces male flowers, which is used to maintain gynoecious lines.

• Melon: CmACS-7 (an ethylene-synthesis gene) acts in carpel primordia to arrest stamens. CmWIP1, a transcription factor, suppresses carpels to give male flowers. CmACS11 represses CmWIP1.
• Cucumber: the F (female) locus is a duplicated ACC synthase gene (CsACS1G).
• Maize (monoecious): tassel seed and dwarf genes show that jasmonic acid is needed to abort pistils in the tassel, and gibberellin to suppress anthers in the ear.

3. Epigenetic and environmental control

DNA methylation stabilises sex expression; for example, methylation of CmWIP1 in melon gives gynoecy. Short days, low temperature and high nitrogen favour female flowers in cucurbits. Infection by the smut Microbotryum makes female Silene form anthers.

Uses: sex-linked markers identify male and female papaya, date palm and asparagus seedlings early. Gynoecious cucumber lines give early, uniform hybrid seed without emasculation.

(c) Protein synthesis in plants, and protein structure and function

1. Transcription (nucleus)

• RNA polymerase II binds the promoter (TATA box) with general transcription factors. Specific transcription factors (for example, auxin response factors) control when it does so.
• The pre-mRNA receives a 5′ 7-methylguanosine cap, introns are removed by the spliceosome, and a poly(A) tail is added.
• The mature mRNA leaves through nuclear pores.
• Plants also have RNA polymerases IV and V, which make small RNAs for silencing, and separate genomes in plastids and mitochondria with their own bacterial-type transcription.

2. Translation (cytosol, on 80S ribosomes)

• Initiation: the 40S subunit with initiator Met-tRNA binds the cap, scans to the first AUG, and the 60S subunit joins.

Elongation:

• An aminoacyl-tRNA, charged by its specific aminoacyl-tRNA synthetase, enters the A site according to codon–anticodon pairing.
• The rRNA of the large subunit forms the peptide bond (the ribosome is a ribozyme).
• The ribosome translocates, the empty tRNA leaves from the E site, and the chain grows from N- to C-terminus. Several ribosomes read one mRNA as a polysome.
• Termination: a stop codon (UAA, UAG or UGA) is recognised by release factors, and the polypeptide is released.
• Chloroplasts and mitochondria have their own 70S ribosomes. For example, RuBisCO's large subunit (rbcL) is made inside the chloroplast.

3. Folding, targeting and modification

• Chaperones (Hsp70, chaperonins) help folding.

Targeting signals decide the destination:

• A signal peptide leads to the ER, then the Golgi, and then secretion, the vacuole or the membrane (for example seed storage proteins and cell-wall extensins).
• A transit peptide leads to chloroplasts. The small subunit of RuBisCO (rbcS) is made in the cytosol, imported, and assembled with the large subunit.
• Presequences lead to mitochondria, and NLS signals to the nucleus.
• Modifications include glycosylation, phosphorylation (signalling), disulfide bonds, lipid anchors and proteolytic processing.
• Proteins are turned over by ubiquitination and the 26S proteasome, the same system through which auxin, gibberellin and jasmonate signal.

4. Structure of proteins

• Primary: the sequence of amino acids joined by peptide bonds. It determines all higher levels (Anfinsen).
• Secondary: α-helices and β-pleated sheets, held by hydrogen bonds between backbone groups.
• Tertiary: the 3-D fold of one chain, stabilised by hydrophobic interactions, hydrogen bonds, ionic bonds and disulfide bridges, and organised into domains.
• Quaternary: the assembly of several subunits. RuBisCO, for example, is L₈S₈, with eight large and eight small subunits.

5. Functions of proteins, with plant examples

• Enzymes: RuBisCO, probably the most abundant protein on Earth; nitrate reductase.
• Storage: gluten proteins (gliadin and glutenin) of wheat, zein of maize, legumin and vicilin of legumes.
• Structure: extensins and other hydroxyproline-rich glycoproteins of the cell wall; tubulin and actin.
• Transport: aquaporins, the plasma-membrane H⁺-ATPase, sucrose transporters.
• Receptors and signalling: phytochrome, phototropins, TIR1, BRI1.
• Regulation: transcription factors such as MADS-box proteins in flower development.
• Defence: pathogenesis-related proteins, lectins, protease inhibitors.
• Movement: myosin XI drives cytoplasmic streaming.
• Biotechnology: proteins are also engineered for crop improvement. Examples are Bt Cry proteins in cotton, and herbicide-tolerant EPSPS.