Efficient targeting of polyhydroxybutyrate biosynthetic enzymes to plant peroxisomes requires more than three amino acids in the carboxyl-terminal signal

Metabolic engineering of plant peroxisomes for biotechnological purposes typically requires efficient peroxisomal targeting of heterologous proteins. Type 1 peroxisomal targeting signals (PTS1) consist of three uncleaved amino acids (SKL or a conserved variant) at the carboxyl terminus and direct nuclear-encoded proteins into the peroxisomes of eukaryotic cells. PTS1 fusion with a heterologous protein results in peroxisomal targeting of that protein, but the minimal length of PTS1 required for efficient targeting in plants is vague. Here, we determine short effective PTS1 sequences derived from plant peroxisomal proteins to target four heterologous proteins, namely the green fluorescent protein (GFP) and the three enzymes required for polyhydroxybutyrate (PHB) production, PhaA, PhaB and PhaC, each fused to the C-terminus of GFP. Transient expression analysis in leaf cells of Saccharum Sp. (Sugarcane interspecific hybrids) indicated that a three amino acid (ARL) PTS1 effectively targeted only GFP and PhaB to peroxisomes. The same signal was not Sufficient to target PhaA and only inefficiently targeted PhaC. An alternative, prototypic three amino acid (SKL) PTS1 was also insufficient to target PhaA and inefficient in targeting PhaC, whilst a six amino acid (RAVARL) PTS1 efficiently targeted both of these enzymes. This study highlights the need for more than a three amino acid PTS1 to target some heterologous proteins to plant peroxisomes. Crown Copyright (C) 2009 Published by Elsevier GmbH. All rights reserved

A proteomic view into infection of greyback canegrubs (Dermolepida albohirtum) by Metarhizium anisopliae

Metarhizium anisopliae is a naturally occurring cosmopolitan fungus infecting greyback canegrubs (Dermolepida albohirtum). The main molecular factors involved in the complex interactions occurring between the greyback canegrubs and M. anisopliae (FI-1045) were investigated by comparing the proteomes of healthy canegrubs, canegrubs infected with Metarhizium and fungus only. Differentially expressed proteins from the infected canegrubs were subjected to mass spectrometry to search for pathogenicity related proteins. Immune-related proteins of canegrubs identified in this study include cytoskeletal proteins (actin), cell communication proteins, proteases and peptidases. Fungal proteins identified include metalloproteins, acyl-CoA, cyclin proteins and chorismate mutase. Comparative proteome analysis provided a view into the cellular reactions triggered in the canegrub in response to the fungal infection at the onset of biological control

The sugarcane genome challenges: Strategies for sequencing a highly complex genome

Sugarcane cultivars derive from interspecific hybrids obtained by crossing Saccharum officinarum and Saccharum spontaneum and provide feedstock used worldwide for sugar and biofuel production. The importance of sugarcane as a bioenergy feedstock has increased interest in the generation of new cultivars optimised for energy production. Cultivar improvement has relied largely on traditional breeding methods, which may be limited by the complexity of inheritance in interspecific polyploid hybrids, and the time-consuming process of selection of plants with desired agronomic traits. In this sense, molecular genetics can assist in the process of developing improved cultivars by generating molecular markers that can be used in the breeding process or by introducing new genes into the sugarcane genome. For meeting each of these, and additional goals, biotechnologists would benefit from a reference genome sequence of a sugarcane cultivar. The sugarcane genome poses challenges that have not been addressed in any prior sequencing project, due to its highly polyploid and aneuploid genome structure with a complete set of homeologous genes predicted to range from 10 to 12 copies (alleles) and to include representatives from each of two different species. Although sugarcane's monoploid genome is about 1 Gb, its highly polymorphic nature represents another significant challenge for obtaining a genuine assembled monoploid genome. With a rich resource of expressed-sequence tag (EST) data in the public domain, the present article describes tools and strategies that may aid in the generation of a reference genome sequence

Nitrogen management guidelines for sugarcane production in Australia: can these be modified for wet tropical conditions using seasonal climate forecasting?

Sugarcane is a highly valuable crop grown in tropical and subtropical climates worldwide primarily for the production of sucrose-based products. The Australian sugarcane industry is located in close proximity to sensitive environments and the apparent declining health of the Great Barrier Reef has been linked to damaging levels of land-based pollutants entering reef waters as a result of sugarcane cultivation undertaken in adjacent catchments. Unprecedented environmental scrutiny of N fertiliser application rates is necessitating improved N fertiliser management strategies in sugarcane.Over time the focus of N fertiliser management has shifted from maximising production to optimising profitability and most recently to improved environmental sustainability. However, current N calculations are limited in their ability to match N fertiliser inputs to forthcoming crop requirements. Seasonal climate forecasts are being used to improve decision-making capabilities across different sectors of the sugarcane value chain. Climate is a key driver of crop growth, N demand and N loss processes, but climate forecasts are not being used to guide N management strategies. Seasonal climate forecasts could be used to develop N management strategies for 'wet' and 'dry' years by guiding application rate, timing and/or frequency of N inputs and the benefit of using alternative forms of N fertiliser. The use of seasonal climate forecasts may allow more environmentally sensitive yet profitable N management strategies to be developed for the Australian sugarcane industry