title: The Biology and Biochemistry of PhoH2 proteins

authors: Andrews, Emma Sophie Vout

subjects: Mycobacteria

abstract: PhoH2 proteins are found in a diverse range of organisms. To date little is known about these proteins and the role they play in the organisms in which they reside. PhoH2 is a PIN PhoH domain fusion, and these proteins are currently annotated as having unknown function and are described as PhoH like. PhoH domains are thought to be ATPases and all characterised PIN domain proteins are RNases. Most efforts have focussed on determining the role of PIN domain proteins that comprise the toxic component (VapC) of VapBC toxin antitoxin systems, in which the PIN domain is coexpressed as part of an operon with an inhibitor (VapB). In the remaining cases where PIN domain proteins can be found such as PIN PhoH domain fusions, these cases remain unexamined. This thesis describes the biological and biochemical characterisation of the PIN PhoH protein, PhoH2 from Mycobacterium tuberculosis and Mycobacterium smegmatis, along with a preliminary structural characterisation of a thermophilic PhoH2 protein homologue. 

The phoH2 gene from both mycobacterial organisms was found to be expressed as part of a long mRNA transcript. Examination of these transcripts revealed possible alternative 5’ start sites out of frame with the phoH2 gene. For protein overexpression, and ‘normal’ growth and colony formation with conditional overexpression, phoH2 from M. tuberculosis required 152 bp of the 5’ sequence directly upstream of the annotated phoH2 gene (annotated here as phoH2alt). PhoH2 proteins: PhoH2alt MTB, PhoH2alt MSMEG and PhoH2MSMEG show ATP/Mg²⁺ dependent, sequence specific RNA unwinding and cleavage. The sequence (A C) (A/U) (A/U) (G/C) U was deduced as a substrate for PhoH2, and PhoH2alt MTB also demonstrated unwinding and cleavage activity on its upstream ~152 base RNA transcript, suggesting a potential autoregulatory mechanism. Structural analysis of a thermophilic PhoH2 protein homologue has provided preliminary crystallographic data which along with electron microscopy suggest a ring like hexameric PhoH2 oligomer.

text: The Biology and Biochemistry of PhoH2 proteins PhoH2 proteins are found in a diverse range of organisms. To date little is known about these proteins and the role they play in the organisms in which they reside. PhoH2 is a PIN PhoH domain fusion, and these proteins are currently annotated as having unknown function and are described as PhoH like. PhoH domains are thought to be ATPases and all characterised PIN domain proteins are RNases. Most efforts have focussed on determining the role of PIN domain proteins that comprise the toxic component (VapC) of VapBC toxin antitoxin systems, in which the PIN domain is coexpressed as part of an operon with an inhibitor (VapB). In the remaining cases where PIN domain proteins can be found such as PIN PhoH domain fusions, these cases remain unexamined. This thesis describes the biological and biochemical characterisation of the PIN PhoH protein, PhoH2 from Mycobacterium tuberculosis and Mycobacterium smegmatis, along with a preliminary structural characterisation of a thermophilic PhoH2 protein homologue. 

The phoH2 gene from both mycobacterial organisms was found to be expressed as part of a long mRNA transcript. Examination of these transcripts revealed possible alternative 5’ start sites out of frame with the phoH2 gene. For protein overexpression, and ‘normal’ growth and colony formation with conditional overexpression, phoH2 from M. tuberculosis required 152 bp of the 5’ sequence directly upstream of the annotated phoH2 gene (annotated here as phoH2alt). PhoH2 proteins: PhoH2alt MTB, PhoH2alt MSMEG and PhoH2MSMEG show ATP/Mg²⁺ dependent, sequence specific RNA unwinding and cleavage. The sequence (A C) (A/U) (A/U) (G/C) U was deduced as a substrate for PhoH2, and PhoH2alt MTB also demonstrated unwinding and cleavage activity on its upstream ~152 base RNA transcript, suggesting a potential autoregulatory mechanism. Structural analysis of a thermophilic PhoH2 protein homologue has provided preliminary crystallographic data which along with electron microscopy suggest a ring like hexameric PhoH2 oligomer.

year: 2013

title: Stratigraphy and sedimentology of Pliocene limestones in northern Hawke's Bay: The Opoiti, Whakapunake and Tahaenui Limestones

authors: Drinnan, Michele Maree

subjects: limestone

abstract: The Opoiti, Whakapunake and Tahaenui Limestone formations (Opoitian to Waipipian; Pliocene) crop out extensively in northern Hawke’s Bay between Wairoa and Mahia Peninsula where they are encased within mudstone dominated Wairoa Formation. The limestones are shallow cool-water carbonates that formed about tectonically active antiform structures inboard from the convergent subduction plate margin along eastern North Island. Their coarse skeletal fraction is dominated by barnacle plates with common brachiopod, pectinid and oyster remains. The carbonate factory for prolific skeletal production was likely sited in shoal water (30-60 m deep), high energy conditions atop the antiforms, with deposition from carbonate shedding down the flanks of the antiforms and even into the bounding synforms.

The individual limestone units are laterally discontinuous and perceived as large lenses (up to c.3-4 km long by c.2 km wide). The enclosing thick mud-rich (M1-M4) to locally sandy (S1-S4) lithofacies, along with occasional volcaniclastic beds (V1), are here informally recorded as Wairoa Formation A, B, C or D, depending on their stratigraphic position with respect to the three limestone units.

Stratigraphic logging of sections at Nuhaka (Tahaenui/Clonkeen), Mt Moumoukai and on Mahia Peninsula established 12 sedimentary lithofacies based on field texture and composition, namely limestones (L1-L3), sandstones (S1-S4), mudstones (M1-M4), and a volcaniclastite (V1). The lithofacies discriminate well on triangular plots involving carbonate content and insoluble sand, silt and clay grain sizes and typically show vertical similarities between field sites for each limestone, suggestive of similar depositional patterns and controls operating at the different localities.

Opoiti Limestone occurs on Mahia Peninsula and Mt Moumoukai as a bedded, c.30 m thick, moderately dipping (25° W), siliciclastic sand-rich unit with common brachiopods and mudstone clasts (L1, L2, L3) that unconformably overlies late Miocene mudstone (M1). Petrography ranges from a sandy biomicrite to a variably sandy, poorly washed rounded biosparite or bioclastic arenite in which interskeletal space is occupied mainly by microbioclastic micrite with some isopachous sparite. Carbonate content is up to c.75%; siliciclastic grains range from very fine to fine sand size. The Opoiti Limestone is likely a transgressive event deposit (TST), fining upwards into sandstone and mudstone (HST) of Wairoa Formation B, and largely under tectonic control.

Whakapunake Limestone within the field area is restricted to the west coast of Mahia Peninsula where it is c.40 m thick and comprises limestone (L3) units (10- 100 cm thick) interbedded with unique mudclast-bearing shelly sandstone (S4) units (20-100 cm thick). These couplets may be related to storm emplacements although their origin, and that of the mudclasts, remains problematic. The elongated mudclasts possibly mark the positions of original Skolithos/Ophiomorpha-like burrows that have been later modified by seismic shaking. No lower contact was observed. Upwards, the limestone grades via interbeds into shelly sandstone (S2) and mudstone (M1). Petrography ranges from packed biomicrites to poorly washed biosparites, while the sandstone interbeds are typically muddy bioclastic arenites. Cements are isopachous sparite rims and microbioclastic micrite with common siliciclasts. Carbonate contents range up to c.70% with the siliciclastic grains being of medium silt and fine sand size. The Whakapunake Limestone is likely a transgressive deposit (TST) fining upwards into shelly sandstone (TST/HST) of Wairoa Formation C, again overall tectonically driven but with possible superimposed eustatic sea level changes.

Tahaenui Limestone (L1, L3) occurs in discrete outcrops (10-30 m thick) across the full area. At Nuhaka, it unconformably overlies late Miocene mudstone (M1), while the upper contact grades into shelly sandstone (S2). At Moumoukai, it unconformably onlaps the Opoiti Limestone. It is dominated by coarse barnacle plates with interparticle isopachous sparite rims and microbioclastic micrite. Petrographically the limestone ranges from rounded biosparite to packed biomicrite. Carbonate content is up to 90%, with the siliciclastic grains being of fine-medium silt and fine sand size. The Tahaenui Limestone is overall a transgressive deposit (TST) fining upwards into shelly sandstone and mudstone (HST) of Wairoa Formation D, largely under tectonic control.

The Pliocene limestones are economically important as potential subsurface petroleum reservoirs, as a lime resource for agricultural use and as a hard stone source for aggregate. Recommendations of the immediate economic potential for development include local aggregate and fertiliser sources from the Tahaenui Limestone at Nuhaka and Mahia.

text: Stratigraphy and sedimentology of Pliocene limestones in northern Hawke's Bay: The Opoiti, Whakapunake and Tahaenui Limestones The Opoiti, Whakapunake and Tahaenui Limestone formations (Opoitian to Waipipian; Pliocene) crop out extensively in northern Hawke’s Bay between Wairoa and Mahia Peninsula where they are encased within mudstone dominated Wairoa Formation. The limestones are shallow cool-water carbonates that formed about tectonically active antiform structures inboard from the convergent subduction plate margin along eastern North Island. Their coarse skeletal fraction is dominated by barnacle plates with common brachiopod, pectinid and oyster remains. The carbonate factory for prolific skeletal production was likely sited in shoal water (30-60 m deep), high energy conditions atop the antiforms, with deposition from carbonate shedding down the flanks of the antiforms and even into the bounding synforms.

The individual limestone units are laterally discontinuous and perceived as large lenses (up to c.3-4 km long by c.2 km wide). The enclosing thick mud-rich (M1-M4) to locally sandy (S1-S4) lithofacies, along with occasional volcaniclastic beds (V1), are here informally recorded as Wairoa Formation A, B, C or D, depending on their stratigraphic position with respect to the three limestone units.

Stratigraphic logging of sections at Nuhaka (Tahaenui/Clonkeen), Mt Moumoukai and on Mahia Peninsula established 12 sedimentary lithofacies based on field texture and composition, namely limestones (L1-L3), sandstones (S1-S4), mudstones (M1-M4), and a volcaniclastite (V1). The lithofacies discriminate well on triangular plots involving carbonate content and insoluble sand, silt and clay grain sizes and typically show vertical similarities between field sites for each limestone, suggestive of similar depositional patterns and controls operating at the different localities.

Opoiti Limestone occurs on Mahia Peninsula and Mt Moumoukai as a bedded, c.30 m thick, moderately dipping (25° W), siliciclastic sand-rich unit with common brachiopods and mudstone clasts (L1, L2, L3) that unconformably overlies late Miocene mudstone (M1). Petrography ranges from a sandy biomicrite to a variably sandy, poorly washed rounded biosparite or bioclastic arenite in which interskeletal space is occupied mainly by microbioclastic micrite with some isopachous sparite. Carbonate content is up to c.75%; siliciclastic grains range from very fine to fine sand size. The Opoiti Limestone is likely a transgressive event deposit (TST), fining upwards into sandstone and mudstone (HST) of Wairoa Formation B, and largely under tectonic control.

Whakapunake Limestone within the field area is restricted to the west coast of Mahia Peninsula where it is c.40 m thick and comprises limestone (L3) units (10- 100 cm thick) interbedded with unique mudclast-bearing shelly sandstone (S4) units (20-100 cm thick). These couplets may be related to storm emplacements although their origin, and that of the mudclasts, remains problematic. The elongated mudclasts possibly mark the positions of original Skolithos/Ophiomorpha-like burrows that have been later modified by seismic shaking. No lower contact was observed. Upwards, the limestone grades via interbeds into shelly sandstone (S2) and mudstone (M1). Petrography ranges from packed biomicrites to poorly washed biosparites, while the sandstone interbeds are typically muddy bioclastic arenites. Cements are isopachous sparite rims and microbioclastic micrite with common siliciclasts. Carbonate contents range up to c.70% with the siliciclastic grains being of medium silt and fine sand size. The Whakapunake Limestone is likely a transgressive deposit (TST) fining upwards into shelly sandstone (TST/HST) of Wairoa Formation C, again overall tectonically driven but with possible superimposed eustatic sea level changes.

Tahaenui Limestone (L1, L3) occurs in discrete outcrops (10-30 m thick) across the full area. At Nuhaka, it unconformably overlies late Miocene mudstone (M1), while the upper contact grades into shelly sandstone (S2). At Moumoukai, it unconformably onlaps the Opoiti Limestone. It is dominated by coarse barnacle plates with interparticle isopachous sparite rims and microbioclastic micrite. Petrographically the limestone ranges from rounded biosparite to packed biomicrite. Carbonate content is up to 90%, with the siliciclastic grains being of fine-medium silt and fine sand size. The Tahaenui Limestone is overall a transgressive deposit (TST) fining upwards into shelly sandstone and mudstone (HST) of Wairoa Formation D, largely under tectonic control.

The Pliocene limestones are economically important as potential subsurface petroleum reservoirs, as a lime resource for agricultural use and as a hard stone source for aggregate. Recommendations of the immediate economic potential for development include local aggregate and fertiliser sources from the Tahaenui Limestone at Nuhaka and Mahia.

year: 2011

title: Ethnic disparities in breast cancer survival in New Zealand: which factors contribute?

authors: Tin Tin, Sandar

subjects: Breast cancer

abstract: Background: New Zealand has major ethnic disparities in breast cancer survival with Maori (indigenous people) and Pacific women (immigrants or descended from immigrants from Pacific Islands) faring much worse than other ethnic groups. This paper identified underlying factors and assessed their relative contribution to this risk differential. 
Methods: This study involved all women who were diagnosed with primary invasive breast cancer in two health regions, covering about 40% of the national population, between January 2000 and June 2014. Maori and Pacific patients were compared with other ethnic groups in terms of demographics, mode of diagnosis, disease factors and treatment factors. Cox regression modelling was performed with stepwise adjustments, and hazards of excess mortality from breast cancer for Maori and Pacific patients were assessed. 
Results: Of the 13,657 patients who were included in this analysis, 1281 (9.4%) were Maori, and 897 (6.6%) were Pacific women. Compared to other ethnic groups, they were younger, more likely to reside in deprived neighbourhoods and to have co-morbidities, and less likely to be diagnosed through screening and with early stage cancer, to be treated in a private care facility, to receive timely cancer treatment, and to receive breast conserving surgery. They had a higher risk of excess mortality from breast cancer (age and year of diagnosis adjusted hazard ratio: 1.76; 95% CI: 1.51-2.04 for Maori and 1.97; 95% CI: 1.67-2.32 for Pacific women), of which 75% and 99% respectively were explained by baseline differences. The most important contributor was late stage at diagnosis. Other contributors included neighbourhood deprivation, mode of diagnosis, type of health care facility where primary cancer treatment was undertaken and type of loco-regional therapy. 
Conclusions: Late diagnosis, deprivation and differential access to and quality of cancer care services were the key contributors to ethnic disparities in breast cancer survival in New Zealand. Our findings underscore the need for a greater equity focus along the breast cancer care pathway, with an emphasis on improving access to early diagnosis for Maori and Pacific women.

text: Ethnic disparities in breast cancer survival in New Zealand: which factors contribute? Background: New Zealand has major ethnic disparities in breast cancer survival with Maori (indigenous people) and Pacific women (immigrants or descended from immigrants from Pacific Islands) faring much worse than other ethnic groups. This paper identified underlying factors and assessed their relative contribution to this risk differential. 
Methods: This study involved all women who were diagnosed with primary invasive breast cancer in two health regions, covering about 40% of the national population, between January 2000 and June 2014. Maori and Pacific patients were compared with other ethnic groups in terms of demographics, mode of diagnosis, disease factors and treatment factors. Cox regression modelling was performed with stepwise adjustments, and hazards of excess mortality from breast cancer for Maori and Pacific patients were assessed. 
Results: Of the 13,657 patients who were included in this analysis, 1281 (9.4%) were Maori, and 897 (6.6%) were Pacific women. Compared to other ethnic groups, they were younger, more likely to reside in deprived neighbourhoods and to have co-morbidities, and less likely to be diagnosed through screening and with early stage cancer, to be treated in a private care facility, to receive timely cancer treatment, and to receive breast conserving surgery. They had a higher risk of excess mortality from breast cancer (age and year of diagnosis adjusted hazard ratio: 1.76; 95% CI: 1.51-2.04 for Maori and 1.97; 95% CI: 1.67-2.32 for Pacific women), of which 75% and 99% respectively were explained by baseline differences. The most important contributor was late stage at diagnosis. Other contributors included neighbourhood deprivation, mode of diagnosis, type of health care facility where primary cancer treatment was undertaken and type of loco-regional therapy. 
Conclusions: Late diagnosis, deprivation and differential access to and quality of cancer care services were the key contributors to ethnic disparities in breast cancer survival in New Zealand. Our findings underscore the need for a greater equity focus along the breast cancer care pathway, with an emphasis on improving access to early diagnosis for Maori and Pacific women.

year: 2018